Needle assembly for aspirating and dispensing from sealed containers

ABSTRACT

A needle assembly has at least one guide member supported by a first body for sliding movement between an extended position and a retracted position. A second body is attached to the guide member. A biasing member biases the guide member to the extended position. A liquid handling needle is coupled to the first body so sliding movement of the guide member toward the retracted position retracts the second body relative to the liquid handling needle. The second body is adapted to contact a sealed receptacle. The biasing member urges the guide member toward said extended position during insertion and during said removal of the needle from the receptacle whereby the second body remains in contact with the sealed receptacle as the guide member moves from its extended position to its retracted position during said insertion and from its retracted position to its extended position during said removal.

CROSS-REFERENCE TO A RELATED PATENT APPLICATION

This application is a continuation of U.S. application Ser. No.11/131,277 (now U.S. Pat. No. 7,549,978), which is a divisional of U.S.application Ser. No. 10/156,245 (now U.S. Pat. No. 6,939,515), whichclaims the benefit of U.S. provisional application No. 60/311,332. Thedisclosure of each of the foregoing patents and applications is herebyincorporated by reference.

TECHNICAL FIELD OF THE INVENTION

The present invention relates to the field of research forpre-formulations or polymorphs. More particularly, the present inventionis directed toward apparatus and methods for performing parallelsynthesis and screening of salts and polymorphic forms of drugcandidates.

BACKGROUND OF THE INVENTION

Combinatorial chemistry has revolutionized the process of drugdiscovery. See, for example, 29 Ace. Chem. Res. 1-170 (1996); 97 Chem.Rev. 349-509 (1997); S. Borman, Chem. Eng. News 43-62 (Feb. 24, 1997);A. M. Thayer, Chem. Eng. News 57-64 (Feb. 12, 1996); N. Terret, 1 DrugDiscovery Today 402 (1996)). Although combinatorial chemistry has to agreat extent eliminated the bottleneck in drug discovery, otherbottlenecks have emerged in getting a drug to market. One suchbottleneck is the selection of salts of active pharmaceuticalingredients in such drugs. Another is the identification of polymorphsand pseudo-polymorphs of drug candidates.

A salt of a compound often has characteristics that are desirable for adrug candidate, including increased water solubility and a highermelting point than the compound itself. Further, different salts of adrug candidate may have disparate and discrete physical properties fromone another. For instance, different salts of a compound may havedifferent melting points or solubilities, or may crystallize indifferent forms and/or under different conditions. Traditional saltselection for a drug candidate requires mixing (e.g., sometimes referredto as synthesizing or formulating) a number of different salts of acompound, recrystallizing the salts under a number of differentconditions to generate a crystalline form, and then characterizing thesalt. This process is time consuming because it has to be reiterated anumber of times to identify salts with desirable characteristics.

Not only do different salts of a drug candidate have differentproperties, different polymorphs of the salt or of the neutral compoundmay also have different physical characteristics. As is known in thepharmaceutical industry, the polymorphic state of an activepharmaceutical ingredient can change the biological profile of the drug.An industry journal published an entire special issue on this topic,Organic Process Research & Development (Vol. 4, No. 5, 2000 and inparticular pp. 370-435), with the issue pointing out, inter alia, thatpolymorphism and crystallization issues affect many industries as wellas pharmaceutical compounds, including explosives, color chemicals andfood additives.

Traditional polymorph characterization requires recrystallizing aneutral drug candidate or a drug candidate salt, characterizing thecrystals, and comparing the crystals to known forms to identifypolymorphs. These steps must be reiterated a large number of times inorder to identify all of the polymorphs of a given neutral compound ordrug candidate salt. Thus, although characterization of polymorphs isadvantageous and, in some cases, necessary, the traditional methods ofidentifying and isolating polymorphs can be tedious. Crystallizing newpolymorphs often requires hundreds to thousands of experiments thatanalyze the effects of varying critical parameters such as temperature,solvent and solvent mixtures, mixing time, cooling rates, stirringrates, and concentrations and methods and process for precipitation,cooling, evaporation, slurry, and thermo-cycling.

One reference in the special issue of Organic Process Research &Development discloses the use of a certain technique for the screeningof potential salts of pharmaceutically active compounds. Bastin et al.“Salt Selection and Optimisation Procedures for Pharmaceutical NewChemical Entities”, Organic Process Research & Development 2000, 4,427-435, incorporated herein by reference. The paper discloses a librarydesign for an array of different salts in different solvents. While thisreference discloses a start at speeding up the pre-formulation process,it fails to follow through with screening in parallel or with highthroughput research into crystallographic polymorphs.

In addition, several published patent applications in the area of highthroughput or combinatorial materials science disclose a process inwhich the materials created in the process can be screened on the sameplate in which they are synthesized. For example, WO 99/59716 disclosesand claims creating solids on a removable reactor base plate and thenperforming X-RAY analysis of the solids. WO 01/34290 and WO 01/34291reportedly relate to a “work station” that employs an array that can betransferred between preparing, screening and characterization stationswithout requiring sample handling, preparation or transfer steps. WO96/11878 also discloses parallel crystallization and screening ofmaterials on a substrate.

WO 01/51919 also reportedly relates to a high throughput method forformation, identification and analysis of diverse solid-forms; however,the methods in this application are extremely broad and vague, such thatthe publication serves merely to identify many problems withoutproviding a solution beyond suggesting high throughput methods. Otherpublications reportedly disclose methods of analyzing polymorphs. Forinstance, WO 01/82659 reports a method of using X-ray diffraction toscreen polymorphs. The publication reports that one can compare theX-ray diffraction pattern acquired for a polymorph and compare it withthe X-ray diffraction patterns of known polymorphs of a compound.However, the publication does not disclose methods for rapidlygenerating the polymorph samples or for using the polymorph comparisonsin drug discovery.

Given the rapid process of drug discovery in the pharmaceutical industrythrough combinatorial chemistry, a need generally exists in industry fora combinatorial or high throughput method and apparatus for theresearch, discovery and development of polymorphs formed by drugcandidates. However, despite the cited work, a process for thesystematic high throughput research of pre-formulations has not beendirectly disclosed.

SUMMARY OF THE INVENTION

The present invention addresses this problem by providing a universalsystem that addresses the need to characterize drug candidates. Thesystem provides, inter alia, generation of libraries, salt selection,polymorph characterization, and other high throughput methods foridentifying and characterizing physical properties of drug candidates,using a variety of reacting and screening options.

Specifically, the invention provides methods, systems and apparatus forperforming combinatorial or high throughput preparing, screening andcharacterization of drug candidate salts and/or crystalline structures(e.g., polymorphs) of drug candidates. These methods, systems andapparatus decrease the time needed to find a suitable form of activeingredient for formulation and allow for additional forms of activepharmaceutical ingredients to be discovered, which may allow foradditional patent coverage, a decreased risk of unwanted polymorphsappearing in later stages of pharmaceutical development or ofcompetitors discovering a related form. In addition, the novel apparatusand methods disclosed herein allow for multiple different drugcandidates to be formulated, crystallized and characterized in parallel,thereby creating a high throughput methodology for pharmaceuticalresearch organizations and others.

In one aspect, the invention provides a workflow that enhances theprocess of identifying and characterizing potential activepharmaceutical ingredients (API) from a drug candidate. In oneembodiment of the invention, salts of a drug candidate of interest areformulated using high throughput and/or combinatorial methods. The drugcandidate salts are then screened to determine a variety of parametersor properties, which may include, without limitation, solubility,partition coefficient (log P), crystallinity, hygroscopicity, Ramanspectral pattern, X-ray diffraction (XRD) pattern and melting point. Thedata obtained from the screening are then analyzed to identify suitabledrug candidate salts. In a preferred embodiment, the formulation,screening and analysis are automated. Further, the workflow may beperformed using the apparatus described herein. In another embodiment,the analysis is performed as the data are generated so that suitablesalts may be rapidly identified. The workflow may be terminated after asuitable salt is selected. In a preferred embodiment, the suitable saltis then subjected to polymorph formulation, characterization andanalysis.

In another aspect, the invention uses high throughput and combinatorialmethods to crystallize, characterize and analyze polymorphs and/orpseudo-polymorphs of a drug candidate of interest. Generating andanalyzing polymorphs may follow directly after salt selection or may beperformed using an existing drug candidate. The drug candidate may be aneutral, acidic or basic compound, or may be a drug candidate salt. Inone embodiment, the drug candidate is recrystallized under a variety ofconditions using high throughput and/or combinatorial methods. The drugcandidate crystals are then screened to determine a number ofcharacteristics of the crystal, including, without limitation,solubility, log P, crystallinity, melting point, hygroscopicity, crystalmorphology and birefringence, as well as X-ray diffraction, infrared(IR), Near IR and Raman spectroscopy, among others. The data obtainedfrom the screening are then analyzed to identify the crystallinestructures of the recrystallized drug candidate. Polymorphs of arecrystallized drug candidate may then be categorized according to thecrystalline structure of the polymorph. In a preferred embodiment,polymorph recrystallization, screening and analysis are automated, andmay be performed with the apparatus described herein. In anotherpreferred embodiment, the analysis is performed as the data aregenerated so that different polymorphs and the conditions that producedthem may be rapidly identified.

The invention further provides a method for selecting solvents for saltselection or polymorph generation. The invention further providesapparatus for high throughput salt preparation, recrystallization,solubility analysis, Raman and X-ray diffraction spectral analysis, andmelting point determinations. The invention also provides hardware andsoftware for controlling the salt selection and polymorphcharacterization methods of the invention and provides systems forautomated high throughput operation of these methods.

Thus, one aspect of the invention is directed toward a high throughputmethod for preparing and characterizing different salts of a drugcandidate. In one embodiment, a library is provided having a pluralityof library members, wherein each library member comprises at least onedrug candidate, and reacting in parallel each of the library memberswith an acid, base or salt to form different salts (e.g., complex saltsor neutrals) of at least one drug candidate. In one embodiment, eachlibrary member may further comprise a solvent. In a preferredembodiment, the first library is comprised of at least eight members inregions on a first substrate, wherein the at least eight memberscomprises at least one drug candidate in an amount of between 0.05 and50 mg of sample, reacting in parallel each of the at least eight memberswith an acid, base or salt to form different salts of at least one drugcandidate. In another preferred embodiment, the drug candidate ispresent in an amount of less than 10 mg. In a further embodiment, thesalts are produced as crystals in glass microtiter plates by cooling,evaporation, precipitation, slurry, or solvent gradients of aliquots ofhot solutions.

In one embodiment, the drug candidate salts form crystalline structures.The drug candidate salt crystals and the supernatant or mother liquormay be left together or may be separated from each other after the saltreaction step, such that the crystals reside on the substrate, typicallyin regions so that the crystals can be screened individually. The methodfurther provides for screening the crystals to identify new forms whilesaid crystals reside on the substrate, as well as screening thesupernatant or mother liquor for solubility of one or more drugcandidate salts in each of the different solvents or solvent mixtures.

In general, salts are screened for at least two properties using varioustests such as, for example, birefringence, melting point, solubility,hygroscopicity, Raman spectroscopy pattern, crystal morphology, X-raypowder diffraction pattern, infrared, near infrared or any othersuitable test. In another embodiment, the salts are screened for atleast three properties, four properties or five properties. In oneembodiment, the salts are screened for at least birefringence, meltingpoint, solubility, Raman spectroscopy pattern and X-ray powderdiffraction pattern.

Another aspect of the invention is polymorph identification and/orcharacterization of a selected drug candidate. The drug candidate may bea neutral, acidic or basic compound or may be a salt of a drugcandidate. In one embodiment, a library is provided and may include aplurality of members that each contain at least one drug candidate andat least one solvent. The library members are subjected to crystallizingconditions in parallel for each of the plurality of members on asubstrate in different solvents or solvent mixtures. Each of the membersare then screened to identify and/or characterize different crystallinestructures of at least one drug candidate. In one embodiment, thelibrary comprises at least eight members in an amount from 0.05 to 50 mgeach, preferably less than 20 mg each. In a further embodiment, thepolymorphs are produced in glass microtiter plates or other opticallytransmissive substrate by cooling, evaporation, precipitation by ananti-solvent, slurry, or solvent gradients of aliquots of hot solutions.In another embodiment, the polymorph characterization may be performedwith a drug candidate or drug candidate salt without previously havingperformed salt selection.

In a further embodiment, the crystalline structures comprising thecrystals and the supernatant or mother liquor is separated from eachother after the recrystallization step such that the crystals reside ona substrate, typically in regions so that the crystals can be screenedindividually or in parallel. The method further provides for screeningthe crystals for at least crystallinity while the crystals reside on thesubstrate, as well as screening the supernatant or mother liquor forsolubility of the one or more drug candidates in the different solventsor solvent mixtures. Other screening tests can be selected from avariety of tests, but a sufficient number of tests are performed to makea determination of the number of polymorphs and/or to identifypolymorphs or salts thereof that may be suitable for drug formulation.

The crystals may be screened for any physical property that would helpidentify a polymorph. In general, at least two properties are screened.The properties may be birefringence, melting point, solubility,hygroscopicity, IR spectroscopy, Near IR spectroscopy, Ramanspectroscopy, crystal morphology, X-Ray powder diffraction pattern orany other suitable screening method. In another embodiment, the crystalsare screened for at least three properties, four properties or fiveproperties to identify and characterize polymorphs. In one embodiment,the crystals are screened for at least birefringence, melting point,solubility, Raman pattern and X-ray powder diffraction pattern.

In one embodiment, crystals may be screened to identify and characterizepolymorphs by optically imaging glass microtiter plates or anotheroptically transmissive substrate that contains crystals of drugcandidates or salts thereof. Two different optical scanning techniquescan be used: transmission and reflection. Optical transmission occurswhen an optical signal passes through an array of material samples.Optical reflection occurs when the optical signal is reflected by thematerial sample. Either one of these scanning techniques determinewhether there are crystalline solids as well as determine thecharacteristics (size or habit) of any crystals. In one embodiment, thecrystals may be optically imaged between polarization filters to measurebirefringence, (e.g., to assess crystallinity).

In another embodiment, the crystals are analyzed by birefringencemeasurements before and after removal of a supernatant or mother liquorto detect unstable solvates or hydrates of drug candidates or saltsthereof. In a further embodiment, individual wells or regions of theoptically transmissive substrate may be imaged under magnification (withand without crossed polarization plates) to determine crystal habit andsize. In yet another embodiment, scattered light measurements can beused to determine if there is a crystalline structure. In anotherembodiment, the crystalline structures may be characterized by theirspectral properties, including, without limitation, Raman, IR, Near IRor X-ray diffraction spectroscopy to characterize the crystals formed onglass microtiter plates or other substrate. Preferred embodimentsinclude Raman and/or X-ray diffraction spectroscopy. The invention alsoprovides methods of using software to analyze the spectral data toidentify polymorphs and/or solvates or hydrates contained in the arrays.In one embodiment, the optical imaging measurements (e.g., birefringenceor crystal morphology), and the spectral measurements (e.g., Raman andX-ray diffraction) are made on the same samples without transfer ofmaterial from the substrate.

In another embodiment, the crystalline structures are formed on a glasssubstrate and/or in an apparatus that has an optical pathway to theregions or vessels so that optical measurements can be made while thecrystalline structure is formed (sometimes referred to herein “in situ”measurements). The crystalline structures may be characterized by theiroptical or spectral properties, including, without limitation,birefringence, Raman spectral pattern, IR pattern, Near IR pattern,and/or light scattering. In some embodiments, the apparatus describedherein can be used for these in situ measurements.

In another aspect, the melting temperature of the crystals may bedetermined. The method involves the steps of heating glass microtiterplates or other substrate comprising drug candidates crystallized underdifferent crystallization conditions while making birefringence,scattering, other optical measurements to determine melting points,and/or other phase transitions including but not limited to loss ofmolecules of solvent or water from solvated crystals. In a preferredembodiment, the melting point of the crystals is determined using aparallel melting point apparatus.

In another aspect, the invention is directed toward making multiplecopies of the arrays by “daughtering” from a parent set of solutions.Thus, in one embodiment, the invention is directed toward a method fortesting drug candidates using daughter libraries, comprising forming alibrary comprised of a plurality members in regions on a firstsubstrate, wherein each of the members comprises at least one drugcandidate and a different solvent or solvent mixture, daughtering themembers to a plurality of second substrates to form a plurality ofdaughter libraries; and subjecting the daughter libraries to differentcrystallization conditions, such as different crystallization methods(e.g., solvent evaporation and precipitation) and/or differentcrystallization parameters (e.g., different temperatures or differentrates of cooling). In another embodiment, daughter libraries areconstructed so that sets of identical crystals can be used indestructive measurements (e.g., melting point, hygroscopicity). Inanother embodiment, daughter libraries are constructed to archivelibraries.

The various assemblies and computer software of this invention may becombined into a flexible workflow to identify and characterizepolymorphs. Similarly, the invention also provides a flexible workflowto identify and characterize different drug candidate salts.

In another aspect, this invention is directed toward a solvent array,wherein the solvent array is chosen to achieve a degree of diversitybased on an analysis of solvent parameters. Physical and chemicalproperties or other characteristics of the solvents are used to groupthe solvents into sets based upon similarities of their physicalproperties or characteristics. The number of physical or chemicalproperties or characteristics and the number of groups into which thesolvents and solvent mixtures are clustered determines the type anddegree of diversity in the solvent array for a given experiment orassay. The diverse solvent arrays may be used in the salt selection andpolymorph generation methods described herein. The invention is alsodirected toward software to implement the use of solvent arrays in themethods and systems of the invention.

In another aspect, the invention is directed toward an apparatus thatmay be used for preparing an array of salts or crystals for screening.The apparatus comprises at least two different assemblies, butpreferably three different assemblies, preferably with interchangeableparts between the different assemblies. In one embodiment, the apparatuscomprises at least two assemblies, one for solubilizing drug candidatesfor recrystallization and/or synthesizing salts of drug candidates (thereactor assembly) and one for crystallizing the compounds (thecrystallization assembly). In another embodiment, the apparatuscomprises three assemblies, including a reactor assembly, a filteringassembly and a crystallizing assembly. Further, one or more of theassemblies are provided separately (e.g., the filtering assembly).

In one aspect, the reactor assembly may include a reactor base having anarray of a plurality of receptacles. Each of the receptacles may beisolated from each other to prevent cross-communication of materialscontained therein. In one embodiment, the reactor base comprises thermalsensors embedded in the reactor base and a thermal block that surroundsthe reactor base. The temperature of the thermal block may be computercontrolled. The reactor assembly may be used in conjunction with adispensing assembly that dispenses at least one object (e.g., ball orstirring flea) into a plurality of receptacles located in the reactorbase.

In another aspect, the filtering assembly includes a reactor basecomprising an array of a plurality of receptacles and a filtrationsubassembly. The filtration subassembly includes a plurality of pairs ofholes, where each pair of holes is associated with a receptacle in thereactor base. One hole may be used for filtering a liquid before it isdeposited into the associated receptacle. The other hole may be used toprovide access to the associated receptacle without having to passthrough a filter. Each pair of holes may isolated from each other toprevent cross-communication of fluid or vapor among other pairs ofholes. Isolation may be accomplished using o-rings that are provided inthe filtration subassembly. In particular, the o-rings may beconstructed to include a large o-ring and a small o-ring. The largeo-ring may surround each pair of holes to provide inter-pair isolation.The small o-ring may surround one of the holes in the pair to provideinter-hole isolation.

In another aspect, the crystallization assembly includes a reactor thatcomprises an optically transmissive substrate. The reactor may include aplurality of through-holes that correspond to regions on the substrate.The through-holes may be sealed such that each through-hole is isolated.In one embodiment, after the crystals form on the substrate, thesubstrate can be removed and subjected to screening test.

In another aspect, the invention is directed toward an apparatus thatcan determine the melting point of a crystal or other solid. In oneembodiment, the melting point apparatus includes a thermal chamber andan image scanning device. An optically transmissive substrate supportingan array of materials (e.g., crystals) may be contained within thethermal chamber. The thermal chamber may heat the substrate at apredetermined rate (e.g., 1° C. per minute) to gradually heat thecrystals on the substrate. As the substrate heats up, the image scanningdevice provides an optical signal to each material sample on thesubstrate and determines whether the crystal, if any, melts or changesphase, or whether the crystal changes composition (i.e., loss of wateror solvent molecules).

The image scanning device may use birefringence imaging, lightscattering, or other optical scanning techniques to determine when acrystal on the substrate melts. Birefringence imaging may beaccomplished using an array of light emitters, polarizer filters andlight detectors. In a preferred embodiment, the image scanning device iscomputer controlled. In addition, the computer may also control the ratein which the temperature of the thermal chamber is increased. Thus, thecombination of temperature and birefringence imaging enables the presentinvention to accurately detect the temperature at which each crystalmelts or changes phase, or whether the crystal changes composition.

In another aspect, the invention is directed toward hardware andsoftware for controlling various apparatuses associated with theinvention. The hardware and software may control the dispensing ofliquid into the reactor assembly, filtering assembly and/orcrystallization assembly. For example, the present invention may be ableto control an automated pipette system to dispense materials into areactor assembly. In one embodiment, the hardware and software can beintegrated with library design software.

Moreover, this invention is directed toward hardware and software forcharacterizing and analyzing one or more physical properties of thecrystals obtained during the polymorph characterization or saltselection methods of the invention. In one embodiment, the softwaregroups the crystals into families of compounds based upon similaritiesin one or more of their physical properties. The physical propertiesthat are used to group the crystals into families may be defined by theuser and/or may be obtained from previous characterization experiments.Further, the different polymorphs or salts may be grouped into familiesbased upon their similarity to one another for one or more physicalproperties of interest within a user-defined deviation. Differentfamilies may be defined by the user based on the same or on differentdeviations. The software enables quick analysis of data obtained fromscreening experiments and provides for a high throughput methods.

Physical properties that may be used to sort polymorphs into familiesinclude, without limitation, crystal shape, melting point and spectralproperties of crystals. In a preferred embodiment, the physical propertythat is used to sort polymorphs into families is a spectrum of thecrystals obtained during the polymorph characterization. In oneembodiment, the spectra may be obtained by Raman, infrared (IR), near-IRor X-ray diffraction spectroscopy, wherein the software sorts spectrainto polymorph families, and in some embodiments this sorting is basedon pattern matching. Thus, different polymorphs are grouped intofamilies based upon their spectral similarities to one another within auser-defined variation, with different families being defined by theuser.

In another aspect, the invention is directed toward a system for makingand characterizing salts from at least one drug candidate. In oneembodiment, the system may include a computer that controls dispensing,heating, and screening of the materials. The computer may controlrobotic equipment to dispense the drug candidate and one or more acidsor bases into the receptacles to provide the mixture. Once the mixtureis in a reactor assembly, a temperature-controlled housing that housesthe reactor assembly may subject the reactor assembly to a predeterminedtemperature to promote dissolution of the drug candidate containedwithin the mixture.

Another aspect this invention is directed toward a system that tests adrug candidate in high throughput mode to make and characterizepolymorphs. In one embodiment, the system includes a reactor assemblyconfigured to contain a plurality of mixtures. The system may alsoinclude a crystallization assembly that is configured to containcrystallized compounds on a substrate. In addition, the system mayinclude a temperature-controlled housing that is configured to containan assembly such as, for example, a reactor assembly, filtrationassembly, or a crystallization assembly and subject the assembly to apredetermined temperature. The system may also include a computer thatcontrols the temperature-controlled housing and obtains data from thescreening devices. The computer analyzes the data to determine if anypolymorphs have formed and categorizes the polymorph into theappropriate family.

Another aspect of the invention is a needle assembly for aspirating aliquid out of or dispensing liquid into a sealed receptacle. Theassembly includes a first body. At least one guide member is supportedby the first body for sliding movement between an extended position anda retracted position. A second body is attached to the at least oneguide member. The second body is relatively farther from the first bodywhen the guide member is in the extended position and relatively closerto the first body when the guide member is in the retracted position. Abiasing member biases the guide member to the extended position. Theassembly also includes a liquid handling needle coupled to the firstbody so sliding movement of the guide member toward the retractedposition retracts the second body relative to the liquid handlingneedle. The second body is adapted to contact the sealed receptacleduring insertion of the liquid handling needle into the receptacle andduring removal of the liquid handling needle from the receptacle. Thebiasing member urges the at least one guide member toward its extendedposition during the insertion and during the removal whereby the secondbody remains in contact with the sealed receptacle as the guide membermoves from its extended position to its retracted position during theinsertion and from its retracted position to its extended positionduring the removal.

A further understanding of the nature and advantages of the presentinvention may be realized by reference to the remaining portions of thespecification and the drawings.

BRIEF DESCRIPTION OF THE DRAWINGS

The above and other objects and advantages of the invention will beapparent upon consideration of the following detailed description, takenin conjunction with the accompanying drawings, in which like referencecharacters refer to like parts throughout, and in which:

FIG. 1 shows an illustrative pre-formulation discovery tool system thatis in accordance with the principles of the present invention;

FIGS. 2A and 2B show an illustrative flow diagram of salt selectionprocess that is in accordance with the principles of the presentinvention;

FIG. 3 shows an illustrative flow diagram of polymorph testing that isin accordance with the principles of the present invention;

FIG. 4 shows an illustrative library that can be modeled in accordancewith the principles of the present invention;

FIG. 5 shows an illustrative flow diagram of a library design process inaccordance with the principles of the present invention;

FIG. 6 shows an illustrative flow diagram that generates a recipe filethat enables automatic control of hardware to synthesize a library inaccordance with the principles of the present invention;

FIG. 7 shows several different crystalline structures that can bedetected with birefringence testing in accordance with the principles ofthe present invention;

FIG. 8 shows an illustrative flow diagram for categorizing samplesaccording to data obtained in screening tests in accordance with theprinciples of the present invention;

FIG. 9 shows a graphical representation of spectral data obtained from aRaman screening device in accordance with the principles of the presentinvention;

FIG. 10 shows a graphical representation of spectral data obtained froma X-ray screening device in accordance with the principles of thepresent invention;

FIG. 11 shows an interactive display screen that includes severalspectral data graphs of crystalline structures discovered in an array ofmaterials in accordance with the principles of the present invention;

FIG. 11A shows an illustrative display screen that enables a user todefine parameters used for correlating data in accordance with theprinciples of the present invention;

FIG. 11B shows an illustrative display screen that enables a user todefine parameters for preforming the correlation of data in accordancewith the principles of the present invention;

FIG. 11C shows an illustrative display screen that enables a userassociate data with particular polymorph families in accordance with theprinciples of the present invention.

FIG. 12 shows an illustrative display screen that shows the spectralgraphs of FIG. 11 in categories in accordance with the principles of thepresent invention;

FIG. 12A shows a flow chart of a process that determines if a materialsample should be selected for additional testing in accordance with theprinciples of the present invention;

FIG. 13 shows a cross-sectional view of a reactor assembly in accordancewith the principles of the present invention;

FIG. 14 shows both a three-dimension view and an explodedthree-dimensional view of a ball dispensing assembly in accordance withthe principles of the present invention;

FIG. 15 shows an exploded view of a reactor assembly in accordance withthe principles of the present invention;

FIG. 16 shows an exploded view of a filtration assembly in accordancewith the principles of the present invention;

FIG. 17 shows an exploded view of a filter subassembly of FIG. 16 inaccordance with the principles of the present invention;

FIG. 18 shows an embodiment of o-ring sheets that may be used in thefilter subassembly of FIG. 17 in accordance with the principles of thepresent invention;

FIG. 19 shows an embodiment of filter disk that can be used inconjunction with the filter assembly of FIG. 16 in accordance with theprinciples of the present invention;

FIGS. 20A and 20B show how a mixture is filtered using the filterassembly of FIG. 16 in accordance with the principles of the presentinvention;

FIG. 20C shows how a filtrate is extracted from the filter assembly ofFIG. 16 in accordance with the principles of the present invention;

FIG. 21 illustrates an exploded view of a crystallization assembly inaccordance with the principles of the present invention;

FIG. 21A illustrates an exploded view of an alternative embodiment of acrystallization assembly in accordance with the principles of thepresent invention;

FIG. 21B illustrates a two-dimensional array of o-rings that are used inconjunction with the crystallization assembly of FIG. 21A in accordancewith the principles of the present invention;

FIG. 21C illustrates a partial cross-sectional view of thecrystallization assembly of FIG. 21A which in accordance with theprinciples of the present invention;

FIG. 22 illustrates a partial cross-sectional view of crystallizationassembly of FIG. 21 that is in accordance with the principles of thepresent invention;

FIG. 22A shows an venting needle assembly in an expanded position inaccordance with the principles of the present invention;

FIG. 22B show a venting needle assembly in a compressed position inaccordance with the principles of the present invention;

FIG. 23 shows a process platform that has a liquid handling robot thatcan dispense and aspirate fluid from assemblies positioned on theplatform in accordance with the principles of the present invention;

FIG. 24 shows perspective top and bottom views of atemperature-controlled housing in accordance with the principles of thepresent invention;

FIG. 25 shows a thermal control chamber in accordance with theprinciples of the present invention;

FIG. 26 shows a three dimensional view of fluid channels of thetemperature-controlled housing of FIG. 25 in accordance with theprinciples of the present invention;

FIG. 27 shows a cross-sectional view of the temperature-controlledhousing that includes a crystallization assembly contained within thehousing in accordance with the principles of the present invention;

FIG. 28 shows an apparatus that simultaneously performs melting pointand birefringence or light scattering testing in accordance with theprinciples of the present invention;

FIG. 28A shows an isometric view of the apparatus of FIG. 28 inaccordance with the principles of the present invention;

FIG. 28B shows a cross-sectional view taken along the line 28-28 of theapparatus in FIG. 28A in accordance with the principles of the presentinvention;

FIGS. 29A, 29B and 29C show Table 3, which lists solvents useful for aprocess of this invention, including certain physical properties of thesolvents that may be used in solvent selection that is in accordancewith the principles of the present invention;

FIG. 30 shows an apparatus for performing in-situ measurement inaccordance with the principles of the present invention; and

FIG. 31 shows an exemplary crystallization workflow using the methodsand apparatus of the invention.

DETAILED DESCRIPTION OF THE INVENTION

Scientific and technical terms used in connection with the presentinvention shall have the meanings that are commonly understood by thoseof ordinary skill in the art with the supplemental definitions foundherein, which are not intended to be contrary to the generally accepteddefinitions. Further, unless otherwise required by context, singularterms shall include pluralities and plural terms shall include thesingular.

As discussed herein, an active pharmaceutical ingredient (API) is aspecific compound (a salt or a neutral compound) that has been approvedby the government for use in a pharmaceutical, e.g., it is safe andeffective for a particular indication.

A drug candidate is a precursor to an API. A drug candidate may haveshown efficacy under various assays for activity or safety under varioustoxicity assays. As used herein, a drug candidate is any compound ofinterest for which pre-formulation testing is desired to determine whichform of the compound can or should be prepared. Pre-formulation testingcan include salt selection and/or polymorph characterization andanalysis. The methodology or process described in this patentapplication may be performed on API's and drug candidates. And, as thoseof skill in the art will appreciate, the exact API or drug candidate isnot critical to this invention, but is typically a small molecule (asopposed to a protein) or a salt thereof. The term drug candidate, asused herein, refers to a neutral compound or a salt thereof, unlessotherwise specified.

A polymorph of a compound, as used herein, is a crystal of a compoundthat is able to crystallize in more than one form. Thus, polymorphs of acompound refer to different crystalline forms of the compound. Apolymorph may also be a pseudo-polymorph, which is a crystal of acompound that contains solvent or water molecules (i.e., solvates andhydrates, respectively) and thus differs from a crystal lacking solventor water molecules. As used herein, the term polymorph refers to bothpolymorphs or pseudo-polymorphs, unless otherwise specified.

A “crystalline structure,” as used herein, comprises a crystal and asupernatant (or mother liquor) formed when a solution crystallizes.

The term “solvent” refers to a liquid that is used to dissolve a drugcandidate in the methods described herein. The term “solvent” is alsointended to include mixtures of solvents (i.e., two or more differentliquids that may or may not be miscible).

The term “salt reactant” refers to acids, bases and salts that are usedto produce drug candidate salts. A “salt reactant” may be definedfunctionally as a substance that provides an ion (either a cation or ananion) that forms a salt with a drug candidate. Generally, a “saltreactant” is added in stoichiometric amounts to drug candidate, e.g., anapproximately equal number of salt ions of the salt reactant are addedas there are acidic or basic moieties on the compound of interest.

A “salt” is defined as a compound comprising anions and cations; thatis, there is a proton transfer between the drug candidate and the acidor base. As used herein, a salt complex is a co-crystal of a drugcandidate and a acid or base; that is, there is no proton transferbetween the drug candidate and the acid or base. Unless indicatedotherwise, the term “salt” as used herein includes both salts and saltcomplexes.

The term “crystallization” or “crystallize” refers to the process bywhich crystals form from a liquid solvent, generating a supernatant(sometimes referred to as a mother liquor) above the crystals.Crystallization also refers to a process in which crystals form frommelts or sublimation. As used herein, a “crystal” is a solid in whichthe molecules are held together in a regular repeating internalarrangement.

As used herein, the term “precipitation” or “precipitate” refers to aprocess by which a solid is generated by addition of an anti-solvent toa drug candidate mixture. The solid may be in either crystalline oramorphous form.

The terms “recrystallization” and “recrystallize” are intended to meancrystallization of the drug candidate compounds from a solution, withoutintending to mean that the drug candidate compounds were crystals priorto being dissolved, unless otherwise indicated.

The term “library” as used herein refers to a plurality of experiments,samples or members, wherein the experiments, samples or members may ormay not be physically associated. Thus, a library refers to members on asingle substrate, on multiple substrates or on a portion of a substrate.In general, each member will have some data associated with it, whichmay include, e.g., drug candidate, salt form, solvent identity, spectraldata, melting point, solubility, etc.

For purposes of the present invention, the term drug candidate is usedgenerically to represent material compounds that are being developed andtested in a process. Further, as used herein, the terms “drug candidate”or “drug candidate compound” are synonymous with the term “compound”,unless otherwise indicated.

The present invention systematically enhances the efficiency of thepre-formulation process of drug development. In particular, thepre-formulation process is implemented as a process that generates,characterizes and analyzes material compositions. This process can beperformed in accordance with the present invention by using illustrativepre-formulation discovery tool system 100 shown in FIG. 1.Pre-formulation system 100 may include computer 110, material handlingapparatus 150, material screening device 160, and user interfaceequipment 180. Pre-formulation system 100 may include multiple materialhandling apparatuses 150 and multiple material screening devices 160.Only one each of material handling device 150 and material screeningdevice 160, however, is illustrated in FIG. 1 to avoid complicating thedrawing. Computer 110 is illustrated to be connected to apparatus 150,device 160, and user interface equipment via communication paths 190. Inaddition, apparatus 150 and device 160 are also connected bycommunication path 190.

Computer 110 controls a process that is implemented to perform thepre-formulation process in accordance with the principles of the presentinvention. Several processes can be implemented in pre-formulationsystem 100. For example, a process may be used for creating, analyzingand selecting salts for a particular drug product. Other processes maybe implemented to generate, characterize and analyze differentcrystalline structures (e.g., polymorphs) of a compound. Comprehensiveprocesses may involve a process that starts with library design ofsolvents and ends with identification of a suitable activepharmaceutical ingredient. Such a comprehensive process may, forexample, combine a salt selection process with a polymorph process topre-formulate a desired ingredient. While there are several possibleprocesses that can be developed and used with pre-formulation system110, the present invention illustrates two such embodiments in FIGS. 2and 3.

Computer 110 may include electronic circuitry 112 (e.g., hard-drive,processing memory communications buses, etc.) that handles transmissionof data to, from, and/or between apparatus 150, device 160, and userinterface equipment 180. Electronic circuitry 112 may enable computer110 to perform processes by controlling, for example, apparatus 150 anddevice 160. Computer 110 may initiate processes by responding to userinput from user interface equipment 180. Computer 110 may also provideinformation to the user at user interface equipment 180 with respect todata acquired during operation of the process.

Electronic circuitry 112 may store, retrieve, and distribute informationfrom database 114. Database 114 stores information that enables aprocess to be created and also provides a basis for performing analysison a material composition. For example, database 114 may storeinformation such as method steps, library designs, results of priorprocesses, results of processes in progress, publicly available data,library compositions, record sets, polymorph family data, and othersuitable pre-formulation data. Database 114 can also store informationon material properties of drug candidates and solvents, such asmolecular weight, density, boiling point, etc. Other stored informationcan include recipe files, reagents, solvents, compounds, salts,crystals, polymorphs, all known characteristics and properties of suchmaterials and any other information suitable for a pre-formulationprocess. Database 114 may be updated with new information. The newinformation may be derived from data obtained from a currently activeprocess or by downloading data via user interface equipment 180.

Computer 110 can run software programs that assist control and operationof a process. Software programs may be used to automate pre-determinedportions of the process. For example, software may automate controlapparatus 150 in preparing library compositions. An illustrative exampleof such software is Impressionist™ software sold by Symyx Technology,Inc. of Santa Clara, Calif. Impressionist™ is described in WO 00/67086,published Nov. 9, 2000, which hereby incorporated by reference in itsentirety. Other software programs may provide a comprehensive computergenerated library that provide the process with a template for preparingvarious material compositions. A computer generated libraryadvantageously eliminates the time consuming task of manuallydetermining all the permutations and combinations of materials that canbe compared under a given set of constraints. An illustrative example oflibrary design software is sold as Library Studio® of SymyxTechnologies, Inc. of Santa Clara, Calif. Library Studio® is describedin WO 00/23921, published Apr. 27, 2000, which hereby incorporated byreference in its entirety. Persons skilled in the art will appreciatethat several software programs may be implemented on computer 110. Forexample, Epoch™ software sold by Symyx Technologies, Inc. of SantaClara, Calif., may be used to control and acquire data from instrumentssuch as apparatus 150 and screening device 160. Epoch™ is described inWO 01/79949, published Oct. 25, 2001, which hereby incorporated byreference in its entirety.

Material handling apparatus 150 may provide assemblies that prepare,filter, and/or crystallize salts or polymorphs of a particular drugcandidate in accordance with the principles of the present invention.Material handling apparatus 150 may be controlled by computer 110 toautomatically mix specific quantities of material (e.g., drugcandidates, reagents, etc.) to form a material composition. Thematerials may be mixed in accordance with library designs produced bycomputer 110 (e.g., with library design software). When the materialsare mixed, they may be analyzed by material screening device 160 todetermine various properties such as solubility, crystallinity, meltingpoint temperature, etc.

If desired, material handling apparatus 150 may also be controlled bycomputer 110 to automate a crystallization process. For example,computer 110 may instruct apparatus 150 to subject its resident materialcompositions to a precipitation process that causes crystallization.After the crystallization process is complete, the material compositionsare tested for at least for crystallinity by screening device 160. Itshould be noted that above apparatus 150 discussion is not intended tobe exhaustive, rather a detailed discussion of material handlingapparatus 150 is discussed below in conjunction with FIGS. 13-30.

Automatic control of material handling apparatus 150 enables the presentinvention to prepare several libraries or members. Moreover, automatedcontrol reduces possible errors that can be caused by manual control. Inaddition, automated control may enable the present invention to preparerelatively small sample sizes (e.g., ranging between nanoliter tomilliliter sizes). This advantageously provides high throughputpreparation and testing of the material compositions.

Material screening device 160 analyzes materials provided on materialhandling apparatus 150 and provides data to computer 110 based on thatanalysis. The analysis of material compositions may also be automatedand controlled by computer 110. Material screening device 160 mayinclude a station such as, for example, a solubility testing station, abirefringence station, spectroscopy stations (e.g., Raman, infrared,X-ray), a melting point station, an electromagnetic signal absorption(e.g., UV-Vis absorption) station, partition coefficient (log P)station, hygroscopicity station, and other suitable devices. One or moreof these stations may provide data that enables computer 110 todetermine the quality and characteristics of a specific material. Forexample, computer 110 may determine the quality of a salt produced basedon the characteristics measured by material screening device 160. Inanother example, material screening device 160 may provide data on thecrystal structure of a material to computer 110. A detailed descriptionof several material screening devices 160 is described below.

User interface equipment 180 enables a user to input commands tocomputer 180 via input device 182. Input device 182 may be any suitabledevice such as, for example, a conventional keyboard, a wirelesskeyboard, a mouse, a touch pad, a trackball, a voice activated console,or any combination of such devices. Input device 182 may enable a userto enter commands to perform drug selection, library building,screening, etc. If desired, input device 182 may enable a user tocontrol material handling apparatus 150 and material screening device160. Using input device 182, for example, a user may calibrate apparatus150 prior to use in a process. In another example, a user may manuallycontrol material screening device 160 to measure materials formulated inmaterial handling apparatus 150. A user may monitor processes operatingon pre-formulation system 100 on display device 184. Display device 184may be a computer monitor, a television, a flat panel display, a liquidcrystal display, a cathode-ray tube (CRT), or any other suitable displaydevice.

Communication paths 190 may be any suitable communications path such as,for example, a cable link, a hard-wired link, a fiber-optic link, aninfrared link, a ribbon-wire link, a blue-tooth link, an analogcommunications link, a digital communications link, or any combinationof such links. Communications paths 190 are configured to enable datatransfer between computer 110, apparatus 150, device 160, and userinterface equipment 180. Communications path 190 may also enable datatransfer between apparatus 150 and device 160.

Various processes can be implemented on pre-formulation system 100.Processes may include several steps or stages to achieve a desiredresult (e.g., salt selection, polymorph testing).

Many neutral pharmaceutically active compounds contain functional groupssuch as amines or carboxylic acids that can react with acids or bases toform salts. In general, salts tend to be more water soluble and havehigher melting points than the corresponding neutral compounds. Saltselection for a drug candidate is not necessary if the neutral compoundhas suitable properties. However, salts of drug candidates often havedesirable properties that are useful for formulation or bioavailability.Desirable properties of a salt compared to a neutral compound mayinclude the ability of the salt to form crystals that are produced moreeasily or cheaply, crystals that are more stable, more filterable, lesshygroscopic, more soluble in water, or have a higher melting point or amore favorable log P for administration. For instance, forpharmaceutical compounds listed in the Physicians' Desk Reference (2000)that are salts, more than 50% of the neutral compounds of the salts areinsoluble in water (they have a solubility of less than 1 mg/mL). Incontrast, less than 20% of the corresponding salts, i.e., the listedpharmaceutical compounds, are similarly insoluble. Further, more than30% of neutral bases listed in the Physicians' Desk Reference (2000)have a melting point below 120° C., while under 10% of theircorresponding salts have a melting point below 120° C. Having a highermelting point is often desirable for easier formulation.

The selection of a salt for a drug candidate of interest is largelylimited to the number of experiments that can reasonably be performed inthe time allotted. Since the counter-ion affects the physical propertyof the drug candidate salt, different salts of the same drug candidatewill crystallize under different conditions and will have differentphysical properties. Thus, it is necessary to perform a number ofdifferent recrystallization experiments to generate crystals fordifferent drug candidate salts. By having a number of different drugcandidate salts, one can identify those salts that are likely to be themost useful for drug formulation and administration.

Table 1, below, lists common anions for salts and Table 2, below, listscommon cations for salts.

TABLE 1 Cumulative Cumulative %: Anion Count % of Total % of Total No.of Compounds Cl 156 60.9 60.9 SO4 26 10.2 71.1 71%: 2 Mesylate 11 4.375.4 Br 10 3.9 79.3 79%: 4 Tartrate 9 3.5 82.8 Citrate 8 3.1 85.9Maleate 7 2.7 88.7 Acetate 4 1.6 90.2 90%: 8 Besylate 4 1.6 91.8 NO3 31.2 93.0 PO4 3 1.2 94.1 Fumerate 3 1.2 95.3  95%: 12 Succinate 3 1.296.5 Benzoate 1 0.4 96.9 Gluconate 1 0.4 97.3 Glucoronate 1 0.4 97.7Lactate 1 0.4 98.0 methylsulfate 1 0.4 98.4 Oleate 1 0.4 98.8 Napsylate1 0.4 99.2  99%: 20 Tannate 1 0.4 99.6 Xinafoate 1 0.4 100.0 Total 256100.0

TABLE 2 Cumulative Cumulative %: Cation Count % of Total % of Total No.of Compounds Sodium 57 76.0 76.0 Potassium 5 6.7 82.7 83: 2 Calcium 45.3 88.0 Magnesium 1 1.3 89.3 89: 4 Ammonium 1 1.3 90.7 Tromethamine 11.3 92.0 t-Butylamine 1 1.3 93.3 Piperazine 1 1.3 94.7 95: 8 Silver 11.3 96.0 Zinc 1 1.3 97.3 Lithium 1 1.3 98.7 Gold 1 1.3 100.0 100: 12Total 75

In Tables 1 and 2, the count number and percent of total indicate thenumber of drugs and the percentage of the total that a particular anionor cation was used as part of an active pharmaceutical ingredient, basedon a survey of compounds in use from the Physicians' Desk Reference(2000). The cumulative percent of the ions is shown in the fourthcolumn. Thus, as shown in Table 1, the two most common anions of activeingredients are chlorides and sulfates, accounting for over 70% of thedrugs on the market as of the printing of the 2000 Physician's DeskReference (PDR). This is highlighted further in the fifth column, whichshows that the top two anions account for 71% of drugs on the market atthat time, the top four account for 79%, the top eight 90%, and the toptwelve 95%. As shown in Table 2, the two most common cations of API'sare sodium and potassium, accounting for over 80% of the drugs on themarket at the time. For cations, the top two cations account for 83% ofdrugs on the market at the time, the top four account for 89%, and thetop eight account for 95%. With the combinatorial or high throughputtechniques of this invention, both anionic and cationic salts can bescreened, such that for most drug candidates, most if not all commonlyused pharmaceutically useful salts can be tested.

In some embodiments of this invention, at least two, four or sixdifferent salt forms of one or more drug candidates are produced andscreened for desired properties. In other embodiments, at least eight,more specifically at least 10, at least 12, at least 16, at least 20, orat least 24 different salt forms of one or more drug candidates areproduced and screened for desired properties. In one embodiment, saltreactants comprising the two, four, six or eight most common anions orcations, as shown in Tables 1 and 2, are used for screening. In anotherembodiment, high throughput screening is performed using salt reactantsfor both anions and cations. Thus, salt reactants for the two, four, sixor eight most common anions and cations are used.

The number of solvents that may be used for salt selection may be anynumber desired. In one embodiment, the number of solvents is two four,six, eight, 12, 16 or more. The screening of salt and solventcombinations (discussed below) can be at the rate of at least eight at atime, at least 12 at a time, at least 24 at a time, at least 36 at atime, at least 48 at a time, or at least 96 at a time. Depending on howthe library is designed, there can be different salts in a plurality ofwells; alternatively, there can be different solvents in a plurality ofwells, etc.

In one embodiment, a different salt is present in each row of asubstrate, wherein a row comprises a number of wells. The substrate maybe a microtiter plate or another substrate on which wells may be formed,e.g., using the salt reaction apparatus described herein. Each column,which also comprises a number of wells and is perpendicular to the rowcontaining the salt, contains a different solvent. See, e.g., FIG. 15,which shows an example of a microtiter format containing rows andcolumns. The library is not limited in this manner. This rate of testingallows for rapid discovery of the appropriate salt(s) for furtherinvestigation.

FIGS. 2A and 2B show an illustrative flow diagram of salt selectionprocess 200 which is in accordance with the principles of the presentinvention. Generally, process 200 dispenses a drug candidate compound ina receptacle in an array format (e.g., an 8 by 12 array) and subjectseach drug candidate to various combinations of one or more saltreactants and, optionally, a solvent. In one embodiment, each receptaclecontains at least one drug candidate, a stoichiometric amount of atleast one salt reactant, and, optionally, a solvent. In a preferredembodiment, each receptacle contains one drug candidate, astoichiometric amount of one salt reactant and a solvent. Depending onthe interaction of mixture of the drug candidate, salt reactant andsolvent, a salt may form and precipitate or crystallize. Salts may thenbe screened and analyzed to determine whether the properties andcharacteristics are suitable for a particular drug application.

Processing selected drug candidates or salts for discovery andcharacterization required at least two, but preferably at least threesteps that are performed in a combinatorial or high throughput mode. Onerequired step is dissolution of drug candidates or salts to form asolution. Another required step is crystallization (e.g., byevaporation, cooling or precipitation with an anti-solvent) of drugcandidates or salts from the solution. An optional processing step priorto crystallization may include separating any remaining solids from thesolution by filtration or centrifugation. This separation or filteringstep may be performed to eliminate nucleation sites in the solventprovided to the crystallization step. Process 200 includes thepreparation, crystallization, filtration, and other steps that areperformed for salt selection.

Prior to preparing solutions comprising the drug candidate and the saltreactant, a user may interact with a computer (e.g., computer 110 ofFIG. 1) to generate a model library. At step 210, one or more computergenerated libraries are generated to enable process 200 to preparesubstantially every possible combination of materials that can be mixedwithin a given set of constraints. Thus, when these mixtures areprepared and then crystallized, the present invention can determinewhich mixture provides the best salt. Each individual mixture iscommonly referred to as a library member or member. The computergenerated model library may be generated using a number of parameters(also called constraints) that typically include materials such as drugcandidates, salt reactants, solvents, environmental conditions, reactionparameters, etc. Parameters may be selected by a user or a softwareprogram. Preferably, a user selects one or more drug candidates formodeling in the library and other parameters such as library size (e.g.,a 96 well array). A library member usually comprises a drug candidate, asalt reactant and a solvent. The solvent, referred to as a librarysolvent in method 200, is used for crystallizing the salt produced bythe reaction of the salt reactant and the drug candidate. Computergenerated library designs are preferably modeled by software programsrunning on computer 110 (shown in FIG. 1). The computer generatedlibrary may be based on user input and information available fromdatabase 114. The computer generated library is described in more detailin conjunction with FIGS. 4 and 5.

Persons skilled in the art will appreciate that computer need not beused to prepare a library. A person could manually prepare a libraryusing any suitable method.

At step 212, a drug candidate and a salt reactant are mixed together. Anoptional mixing solvent, which may be different or the same as thelibrary solvent, may be added to the salt reactant and drug candidate.The drug candidates, salt reactants and optional mixing solvents arereacted together to produce drug candidate salts. At optional step 214,after the drug candidate salts have been produced, the drug candidatesalts are isolated. At step 216, the library solvents provided at step210 are added to the drug candidate salts to produce the librarymembers.

In a preferred embodiment, the drug candidate mixture described at step212 is prepared in a reactor assembly. Reactor assemblies are typicallyconstructed in microtiter format (e.g., an 8×12 96-well plate).Microtiter format is particular useful for performing high throughputreaction of materials. However, any other format may be used (e.g., a384 well plate). In addition, this enables process 200 to construct alibrary in reactor assembly 1300 in accordance with the computergenerated library provided at step 210.

At step 212, drug candidates may be mixed with salt reactants accordingto the computer generated library at step 210. The mixture mayoptionally comprise one or more mixing solvents to provide a saltsolution. The drug candidate is typically dispensed (into reactioncontainers) as a solution or a slurry, but it can be also be dispensedas solid. A liquid dispensing device is illustrated in FIG. 23. A soliddispensing device (not shown), for example, is sold as Powdernium byAutoDose of Geneva, Switzerland. Assuming that the drug candidate is ina solution or slurry, the solvent may or may not be driven off acrossthe plate in parallel (such as by blowing nitrogen over the library orwith a solvent evaporator, e.g., Genevac HT-8 (Genevac Inc, ValleyCottage, N.Y. 10989) under reduced pressure or vacuum. If the drugcandidate is dispensed in the reaction solvent, then solvent removal isunnecessary.

After dispensing of the drug candidate, the chosen salt reactant (e.g.,an acid or base in solution, slurry or solid format) is dispensed intothe wells of the array. For anion salts, the corresponding acid is used.For example, hydrochloric acid, sulfuric acid, mesylic acid and bromicacid may be used for chloride, sulfate, mesylate and bromate salts ofcompounds. For cationic salts, one may use the corresponding hydroxideor other base. For instance sodium hydroxide and potassium hydroxide maybe used for sodium and potassium salts. Magnesium and calcium salts ofdrug candidates may be formed by using magnesium or calcium acetate,oxide or carbonate. Amine salts may be formed by mixing the amine ofchoice itself with the drug candidate. In one embodiment, the saltreactant is dispensed automatically into the wells of the array, asdiscussed in more detail below. In another embodiment, the salt reactantis dispensed manually into the wells of the array.

The salt solution is subjected to various conditions to allow the drugcandidate to react with the salt reactant under the conditions imposed.Common reaction conditions are a temperature of about room temperatureor higher with shaking in sealed vials. The temperature may beapproximately 20° C., 30° C., 40° C., 50° C., 60° C., 70° C., 80° C. orhigher. In one embodiment, the temperature is raised to approximately70° C. to allow the drug candidate to react with the acid or base.Depending on the drug candidate and the salt reactant, a reactionbetween the drug candidate and the reaction solvents may or may notoccur.

Individual vials or wells may contain magnetic stirring fleas that aretumbled (e.g., rotated) by a magnetic field or a rotating magnet. Therotating stirring fleas promote mixing of the materials contained withinthe vials or wells. A detailed explanation of a device that providesmagnetic stirring is described in U.S. Pat. No. 6,176,609, which ishereby incorporated by reference in its entirety.

At optional step 214, after each library member is prepared, anyremaining salt reactants and optional solvents are removed from thereaction mixture. This may be done by any method known in the art,including, e.g., methods used for crystallization, such as evaporation,cooling or addition of an anti-solvent.

At step 216, one or more library solvents are added to each well inaccordance with the computer generated library and the drug candidate isdissolved in the solvent.

At optional step 220, after each library member is prepared, an aliquotof the salt solution is taken from each reaction container and filteredby a filtration assembly. Each salt solution is filtered so that a“seedless” salt solution is used for crystallization at step 222. FIGS.16-20 illustrate various filtration apparatuses that provide filtrationin accordance with the principles of the present invention.

If seeding is desired, a seed may be added separately, so that seededrecrystallization is controlled.

At step 222, the filtered salt solutions are crystallized. Saltcrystallization can occur by 1) cooling the solution, 2) precipitatingthe solution by adding an anti-solvent that causes precipitation, 3)evaporating the solution, or 4) slurrying the solution. If saltcrystallization or precipitation occurs on a multi-well plate or asubstrate, such as the universal substrate described herein, the plateor substrate may be screened by various identifying and characterizationdevices. If desired, characterization can be performed whilerecrystallization is occurring. The salt reactants and solvents that maybe used to crystallize or precipitate the salts are described below insolvent selection section.

Each library member is screened to determine if a salt has formed.Screening is performed at step 225 and can be implemented using materialscreening device 160 (shown in FIG. 1). Screening at step 225 may beperformed primarily as a high-throughput screen that quicklycharacterizes each library member. Data obtained during the screeningmay be provided to a computer (e.g., computer 110). For example,screening at step 225 may obtain a sufficient quantity of data on eachlibrary member such that process 200 can be used to quickly analyze thesalt. The apparatus and methods described herein provide severalscreening methods and devices that can determine birefringence, log P,crystallinity, solubility, and melting point of each library member insitu.

At step 230, process 200 can perform a quick preliminary assessment onthe suitability of each salt before additional time consuming tests areperformed. Analysis can include user-defined selection by selectingwhich library members should be explored further. Salt selection canalso be performed, for example, by computer. A computer can run softwareprograms that analyze the data. For example, results of the screeningcan provide information regarding whether the salt formed a solid,whether the solid was amorphous or crystalline, and informationregarding the solubility, log P and melting point of the crystal.

At step 240, process 200 determines if a salt should be selected forfurther testing or if it should be discarded. If none of the saltsproduced in the library are suitable for additional testing (e.g.,screening), the process returns to step 212 so that new librarycompositions can be prepared. The data obtained from these salts,however, is stored on a database and used for future analysis.Preferably, process 200 continues to prepare library compositions basedon computer generated library models generated in step 210. However, ifstep 240 determines that there is at least one suitable salt, processcontinues to step 250.

At step 250 in FIG. 2B, the salt selected in step 240 is resynthesizedin bulk format. This step is only necessary if sufficient quantities ofthe salt selected in step 240 were not produced. In step 250, multiplesamples of the same material composition are deposited in a differentreactor assembly. If desired, additional solvents may be applied to theexisting library compositions to obtain, for example, additional saltcharacteristics. The application of new materials may be based on alibrary modeled at computer 110 (FIG. 1). Then the newly-formulatedlibrary compositions are crystallized to form salt.

At step 255, the salts are screened again to obtain property andcharacteristics data, if more data are required. This screening can beused to provide a more in-depth characterization of the salts thanscreening step 225. Step 255 obtains data that enables process 200 toperform a secondary analysis (shown at step 260). At step 260, asubstantial analysis may be performed using the data provided by step255. At step 270, process 200 determines if the salt should be selectedfor further testing. If the salt is not selected, process 200 may returnto step 212 (FIG. 2A). If the salt is selected, then process 200 hasdiscovered a salt that is suitable for use in another application.Persons skilled in the art will appreciate that one or more salt formsmay be selected by process 200 and it will also be appreciated thatprocess 200 may not produce any salts that are selected based on theparameters set in step 210.

If desired, after a salt is selected process 200 may proceed to step 310of FIG. 3. As described below, FIG. 3 describes polymorph generation andcharacterization based on the selected salt.

Process 200 may be implemented on pre-formulation system 100 to selectsalts that are suitable for drug development. Process 200 may be used asprecursor to another process that performs test on one or more selectedsalts to determine various potential polymorphs of the salt'scrystalline structure. Persons skilled in the art will appreciate thatsteps shown in FIG. 2 are merely exemplary and that additional steps maybe added and some steps may be omitted or modified. For example, a stepcan be added that causes process 200 to prepare and test every librarymember provided by the computer generated library in step 210 even ifone or more suitable salts have been discovered. Further, daughterlibraries may be formed to alter the conditions under which salts may beformed (e.g., temperature or temperature ramp rate) or to provideidentical salts for destructive analyses (e.g., melting point screens).

FIG. 3 shows an illustrative flow diagram of polymorph discovery process300 which is in accordance with the principles of the present invention.Process 300 may be implemented on pre-formulation system 100 independentof other processes or it may be merged with processes such as process200 of FIGS. 2A and 2B.

Process 300 recrystallizes one or more drug candidates or salts (i.e.,subjecting them to different conditions to generate as many polymorphsas possible, preferably substantially every polymorph, for a particulardrug candidate or salt thereof) and screens each of the polymorphs in ahigh-throughput capacity. This enables process 300 to quicklycharacterize and determine those recrystallization conditions that arebest for developing a desired drug ingredient that possesses a suitablecrystalline structure. As used herein, recrystallization andcrystallization conditions refer to those conditions that affectrecrystallization. These conditions include, e.g., temperature, seeding(if present), solvent(s), etc.

Process 300 begins at step 310. At step 310, process 300 is providedwith one or more drug candidate salts (or neutral compounds) forpolymorph testing. The drug candidates can be preselected, for example,by a user via user interface equipment 180 (shown in FIG. 1). Drugcandidates may also be provided by salts selected in process 200 ofFIGS. 2A and 2B. Alternatively, step 310 may be provided with one ormore drug candidates that can be used for polymorph testing.

Once a drug candidate is provided at step 310, the drug candidate mayundergo recrystallization at step 320. If desired, recrystallization mayinvolve several steps to form crystals from a particular drug candidate.For example, after a drug candidate has been selected, it may be mixedwith solvents 322 in a reaction assembly (e.g., reaction assembly 1300of FIG. 13 or reaction assembly 1500 of FIG. 15). The drug candidate andsolvents may be dispensed into a reaction assembly using a liquiddispensing assembly as described for salt selection process 200. Thesolvents may be mixed in accordance with a computer generated library321 to provide as many different crystallization mixtures for aparticular set of constraints. Solvent selection is described furtherbelow.

The number of solvents that may be used for polymorph generation andcharacterization may be any number desired. In one embodiment, thenumber of solvents is two, four, six, eight, 12, 16, 24, 36, 48, 96 ormore. The number of solvents that may be used for polymorph generationand characterization for a single assay can be at the rate of at leasteight to ten at a time, at least 12 to 14 at a time, at least 24 at atime, at least 36 at a time, at least 48 at a time, or at least 96 at atime. The term solvent in this respect means both a single solvent(e.g., heptane or water) as well as combinations of solvents (e.g.,heptane mixed with water), as described herein.

In one embodiment of this invention, at least one drug candidate is usedin a process of generating and characterizing polymorphs. In otherembodiments at least two, three or four drug candidates are used togenerate and characterize polymorphs.

If further desired, recrystallization may include filtering the mixturesto provide “seedless” or “pure” solutions for crystallization. Afiltering step is performed formulation and prior to crystallization.Crystallizing filtered solutions eliminates nucleation sites that canbias formation of crystals during the crystallization step. Filteringmay be accomplished using filtration assembly 1600 of FIG. 16.

Various recrystallization conditions (e.g., temperature, pressure, time,etc.) may be varied to provide various crystal formation. FIGS. 21 and22 illustrate, for example, an apparatus that can be used to crystallizethe mixtures. One advantage of this apparatus (e.g., crystallizationassembly 2100 of FIG. 21) is that crystals may grow, form or re-salt ona removable substrate. This substrate may be provided to one or morescreening devices such that crystals are scanned for their propertiesand characteristics at step 330.

To perform crystallization or recrystallization under a variety ofcrystallization conditions, daughter libraries may be created from drugcandidates, mixtures, or salts thereof. A daughter library is created bytaking one or more aliquots from one or more members in a parent librarycontained within a reactor assembly. A parent library may includemixtures that are prepared at the beginning of recrystallization step320. An aliquot is a definite fraction of a whole.

To perform daughtering, a pipette, operated either manually orautomatically (e.g., robotically), draws a portion of a member from theparent library and dispenses that aliquot into another container (e.g.,crystallization assembly) to provide a daughter library member. Alimited number of members of the parent library may be daughtered or allthe members may be daughtered at least once to create a daughterlibrary. Thus, a daughter library may be smaller than the parent libraryin terms of either mass, volume or moles and/or in terms of the numberof members. Daughtering is performed, for example, to allow for multipleexperiments on identical mixtures, solutions, or samples to avoid havingto recreate the parent library. There is known equipment that canperform daughtering, such as hand pipetters, hand-multichannelpipetters, or robots (such as Matrix or CyberLab or Hydra robots). Anynumber of daughter libraries may be produced, provided that the originallibrary is of sufficient volume. In one embodiment, at least one, two,four, eight or twelve daughter libraries are produced.

After the mixtures from the parent library and daughter libraries arecrystallized, each library member is screened. Step 330 provides primaryscreening that provides data to step 335, which analyzes the data todetermine if any polymorphs have formed.

The crystals may be screened for any physical property that would helpcharacterize and/or identify a polymorph. The crystals may be screenedfor birefringence, melting point, solubility, hygroscopicity, IRpattern, Near IR pattern or Raman pattern, crystal morphology, X-raypowder diffraction pattern or any other suitable screening method todetermine if crystals (or polymorphs) have formed. In general, at leasttwo properties are screened. In one embodiment, Raman pattern, X-raydiffraction pattern, melting point, birefringence and hygroscopicityscreens are performed to adequately characterize the crystallinestructures.

At step 340, process 300 determines whether a polymorph should beselected being produced in bulk for further testing, if sufficientquantities of the drug candidate have not been produced during process300. If the polymorph is unsuitable, the process may revert back to step320 such that the same or different drug candidates can undergorecrystallization. If the polymorph is selected, process 300 may prepareseveral of the same crystals at step 345 so that they can be screenedand characterized at step 350.

At step 350, the screening is more detailed than the screening performedat step 330. The crystals may be screened for at least two properties,three properties, four properties or five properties to identify andcharacterize polymorphs, wherein the properties are determined by, e.g.,birefringence, melting point, solubility, hygroscopicity, IR pattern,Near IR pattern, Raman pattern, crystal morphology or X-ray powderdiffraction pattern or, if sufficient amounts have been produced, singlecrystal X-ray diffraction, thermogravimetric analysis, nuclear magneticresonance or differential scanning calorimetry. In one embodiment, thecrystals are screened for at least birefringence, melting point,solubility, Raman pattern and X-ray powder diffraction pattern.Preferred embodiments include Raman and/or X-ray diffractionspectroscopy. The process may be performed using a solubility testingstation, a birefringence station, spectroscopy stations (e.g., Raman,infrared, X-ray), a melting point station, an electromagnetic signalabsorption (e.g., UV-Vis absorption) station, partition coefficient (logP) station, using the apparatus described herein.

The data obtained in step 350 may provide a substantial quantity ofinformation to enable process 300 to analyze the polymorphs orcrystalline structures (step 360). At step 365, process 300 maydetermine if the polymorph (e.g., neutral drug candidate compound orsalt thereof) is suitable for use as an active pharmaceutical ingredient(step 370). If the polymorph is not suitable, then that particular drugcandidate is not used, at least for the drug application prescribed forprocess 300 (step 375). The data obtained from analysis, however, may bestored for future reference.

As illustrated in FIGS. 2A, 2B, and 3, processes use computer generatedlibraries as a template for preparing library members. For example,libraries containing various combinations of drug candidates, salts,crystals, and other pre-formulation materials may be generated prior tolibrary preparation. FIG. 4 shows an illustrative library 400 that canbe generated in accordance with the principles of the present invention.Library 400 can include any suitable number of library elements 410.Preferably the number of library elements modeled in each library 400 isthe same as the number of library members 410 that can be prepared onhardware (e.g., a substrate). Any number of rows and columns of librarymembers 410 can be created, thus providing flexibility in librarygeneration. This is illustrated by the variable “N”, which indicatesthat any suitable number of rows and columns can be designed in library400. The row may, for example be Industry typically uses a substratethat has 96 wells or vials for formulating or mixing various materials.Therefore, if library 400 is modeled to have, for example, eight rowsand twelve columns (a typical microtiter plate), library elements 410may be readily implemented in practice. Likewise, if a 384 wellsubstrate is being used, then library 400 may be modeled to have, forexample, 16 rows and 24 columns.

Regardless of the size of library 400, each library element 410 includesat least two materials. The materials in each library element 410 candiffer by drug candidates, different known crystal structures of drugcandidates, solvents and salt reactants. FIG. 4 shows a few examplematerials 420 that can be used to model library elements 410. Based onmaterials 420 and other factors, a computer (e.g., computer 110 ofFIG. 1) can model substantially every possible combination of libraryelements that can be practically constructed. For example, libraryelement 411 illustrates one such possible combination that includes adrug candidate, the salt reactant tartaric acid, and heptane. Thus,generating libraries provides the present invention with a foundation toprepare and test several different material compositions. Thisfacilitates salt selection and provides a basis for discoveringpolymorphs for a particular drug candidate.

A large number of solvents are known that can be used inrecrystallization, either for salt selection and/or for polymorphgeneration. Table 3 (shown in FIGS. 29A, 29B and 29C) lists a number ofexemplary solvents along with some of their physical properties. Giventhe large number of solvents that may be used, it is advantageous tocluster solvents into groups based upon certain shared physicalproperties or other shared characteristics of the solvents and then pickat least one solvent from each group to test in recrystallization. Thisprocess ensures that a wide variety of different types of solvents willbe used for recrystallization, which is advantageous for identifyingpolymorphs or desirable crystals of drug candidate salts.

Table 3 shows solvents along with their chemical class, a referencenumber (#) assigned to different classes of solvents (for statisticalsort purposes), molecular weight (MW), density (n), molar volume(V_(m)), melting point (MP) (in ° C.), boiling point (BP) (in ° C.),enthalpy of evaporation (ΔH_(vap)), Hildebrand solubility parameter (δ),dipole moment (μ), log solubility in water (log S), partitioncoefficient (log P), viscosity, index of refraction, pKa (in water anddimethylsulfoxide (DMSO)), dielectric constant (∈) and ionizationpotential (IE) and pK_(a+). Other parameters known to those in the artcould be included, such as, without limitation, the cost in US dollarsper kilogram, cost of disposal or storage, degree of toxicity orenvironmental safety. The description of the solvents and physicalproperties provided herein does not limit the type of solvent or thephysical properties of the solvent that may be used in the saltselection and polymorph workflows.

One or more of the physical properties of the solvents in a solventlibrary may be used to cluster solvents into groups. The solvents in aparticular group will have similar physical properties for the one ormore properties that has been chosen as a criteria. The physicalproperties of the solvents may be one or more of those listed in Table 3or may be other physical properties known in the art. One may use fromone to n properties to cluster the solvents into groups, wherein n isthe total number of physical properties that have been provided for alibrary of solvents. Each solvent may be independently sorted into agroup with other solvents that have the same defined characteristics forall of the selected physical properties. One may use any number ofproperties to cluster solvents, including from four to 20, four to 16 orsix to eight properties to cluster the solvents. In one embodiment, theclass of the solvent (e.g., whether it is an alcohol, ketone, etc.) isnot employed as one of the physical properties used in grouping thesolvents.

For any particular physical property, the selection criteria may bedefined to provide for two or more different subsets. In one embodiment,the selection criteria for single physical property solvents may providefor two, four, six, eight or twelve subsets. For example, one mayprovide four definitions for dividing the solvents based upon onephysical property (e.g., the solvent's density) and provide twodefinitions for dividing the solvents based upon another physicalproperty (e.g., the solvent's dipole moment). The solvents would then beclustered into eight groups based upon the definitions of these twophysical properties.

In a preferred embodiment, the physical properties of a large number ofsolvents are kept in a database. During library design for polymorphgeneration or salt selection, a user can define both the specificphysical properties and the selection criteria for these physicalproperties in order to cluster the solvents into groups based upon theparticular physical properties chosen and the selection criteria forthese physical properties.

One may design solvent groups to form from two groups up to n groups,wherein n is the total number of solvents in the designated solventlibrary. In one embodiment, the number of solvent groups is four to 96groups. In another embodiment, the number of solvent groups is six to64, eight to 48 groups, ten to 40 groups or 12 to 24 groups. In anotherembodiment, the number of solvent groups is eight to 24 groups, 16 to 24groups, 20 to 40 groups or eight to 12 groups. In general, a smallernumber of solvent groups are used for the salt selection workflow ascompared to the polymorph characterization workflow. For salt selection,the number of solvent groups is generally in the range of 16 to 24groups, while for polymorph identification and characterization, thenumber of solvent groups is generally around 20 to 40.

In one embodiment, in order to provide diversity of the types ofsolvents that are used for salt selection or polymorph generation, asolvent library may be clustered into groups of solvents havingparticular similar physical properties or characteristics and at leastone solvent from each group is used in the salt selection or polymorphgeneration. In another embodiment, a number of solvents from a singlegroup may be selected and used in salt selection or in polymorphgeneration. This may be particularly useful at later stages in polymorphcharacterization, to identify potential facile, stable or commercialmethods for generating a particular polymorph for an activepharmaceutical ingredient, or to prepare focused solvent libraries.

Solvent group design may be performed by any method known in the art,including manual design or computer design. Commercially availablecomputer programs can be used, including JMPT™, available from SASInstitute, Inc., Cary, N.C.

In general, solvents are chosen to have a boiling point higher than thetemperature at which the crystallization will be run. Other physicalproperties that are preferred are those in which non-toxic solvents areused. In another embodiment, one or more solvents such as ethanol,water, cyclohexane, propanol, acetonitrile, dioxane, methyl ethylketone, ethyl acetate, isopropyl acetate, propyl acetate or toluene areused as a solvent in the methods described herein.

In one embodiment of a salt selection array, one axis of the array(e.g., the rows of a microtiter plate or other substrate) contains aconstant amount of the drug candidate of interest mixed with a number ofdifferent salt reactants, wherein each row of the array contains adifferent salt reactant. In general, the drug candidate is mixed withone equivalent of the salt reactant (e.g., an acid or base). However, inanother embodiment, the drug candidate may be mixed with 0.5, two, 1.5,three or four equivalents of the salt reactant.

The opposite axis of the array (e.g., the columns of the microtiterplate) contains a number of different solvents, wherein each column ofthe array contains a different solvent. In a preferred embodiment, thesolvents have been clustered as described herein. Thus, each well of thearray contains a different combination of salt reactant and solvent.

The salt reactant and drug candidate may be subjected to conditions inwhich a salt may form prior to addition of the solvent, e.g., byheating, stirring, shaking or any combination thereof. An array ofsolvents may then be added and the presence of crystals or precipitatesof drug candidate salts may be determined. Alternatively, the drugcandidate, salt reactant and solvent may be mixed together prior tosubjecting the array to conditions under which a drug candidate salt andcrystal or precipitate thereof may form. In one embodiment, the arraycan be subjected to salt synthesis and crystallization using one or moreof the apparatus described herein.

In another embodiment, one axis of the array (e.g., the row) containsthe drug candidate of interest mixed with a number of different saltreactants while the opposite axis (e.g., the column) contains solventsor compositions of two or more solvents, wherein each column contains adifferent solvent or solvent composition. In one embodiment, the solventcompositions in the array or part thereof may be differentconcentrations of the two solvents relative to each other. Theconcentrations of the solvents relative to one another may be anyconcentration desired. In one embodiment, an array of solventcompositions (e.g., gradient) for solvents A and B can be expressed inthe following formula:

x*A(100−x)*B=0  (1)

where x is the percentage.

If desired the gradient can be determined using a linear function, anon-linear function, a polynomial function, an exponential function,etc. For instance, one may vary the concentration of water and heptanein this manner. Similarly, an array of solvent compositions using three,four or more different solvents may be produced, wherein the relativeconcentrations of the solvents to each other are varied in the array. Ina preferred embodiment, the solvents in the solvent composition havebeen clustered as described herein. As discussed above, the array canthen be subjected to conditions in which drug candidate salts wouldlikely form crystals.

A single drug candidate (either a salt or a neutral compound) istypically tested for polymorph generation and characterization, althoughtwo, four, six, eight or more may also be characterized. Becausedifferent drug candidate salts do not have to be formed, more solventsare typically used for polymorph generation and characterization than istypical for salt selection. However, except for the larger number ofsolvents that are generally used, solvent group selection for polymorphgeneration and characterization is similar to that for salt selection.Thus, in one embodiment, a library of solvents may be divided into moregroups for recrystallization compared to that for salt selection, whilein another embodiment, more solvents within a single group may be usedto recrystallize a drug candidate salt.

In one embodiment of a polymorph array, each well contains a constantamount of the drug candidate of interest and a different solvent. Thesolvents may be selected by any method known in the art. For example,the solvents may be selected from different industrially importantcategories of solvents, including aromatics, ketones, water,halogenates, alcohols, esters, nitriles, as well as solvent mixturesthat span a wide range of polarity and dielectric constant. In apreferred embodiment, the solvents are selected by the solvent selectionmethod described previously, by dividing a library of solvents intogroups based upon a variety of different physical properties orcharacteristics. More preferably, the solvents are not selected basedupon their chemical class. After the solvent has been added to the drugcandidate, they are mixed under conditions to dissolve the drugcandidate, optionally filtered, and then subjected to conditions inwhich crystals are likely to form.

Using an array of different types of solvents, whether based upon thechemical class or their physical properties, provides a large amount ofinformation regarding which types of solvents recrystallize the drugcandidate. Further, using a wide variety of solvents is likely toprovide a large number of different polymorphs of the crystalline drugcandidate, if they exist. See, e.g., FIG. 7, which shows changes incrystal morphology of a drug candidate after crystallization fromdifferent solvents. These solvents included those that are miscible inheptane and those that are miscible in water. Once one or more solventshave been identified that recrystallize the drug candidate, othersolvents that are of the same chemical class or share physicalproperties or characteristics with the identified solvents may be usedin further experiments. These experiments can be performed to generateand characterize polymorphs of the drug candidate as well as to find thebest recrystallization solvent for the particular drug candidate.

In another embodiment of a polymorph array, the array contains the drugcandidate of interest mixed with compositions of two or more solvents,wherein each well contains a different solvent composition. See, e.g.,Example 3. In one embodiment, the solvent compositions in the array orpart thereof may be different concentrations of the two solventsrelative to each other. The concentrations of the solvents relative toone another may be any concentration desired. In one embodiment, anarray of solvent compositions for solvents A and B may be determinedusing equation 1. Similarly, an array of solvent compositions usingthree, four or more different solvents may be produced, wherein therelative concentrations of the solvents to each other are varied in thearray. In a preferred embodiment, the solvents in the solventcomposition have been clustered as described herein. As discussed above,the array can then be subjected to conditions in which crystals wouldlikely be synthesized, and the presence of crystals and polymorphs canbe determined. One can use different concentrations of solvents todetermine at what concentration a solvent causes the drug candidate tocrystallize. In addition, one can use different concentrations todetermine whether different types of crystals are formed in differentconcentrations of solvent.

In another embodiment, solvent group design may be performed byproviding a database containing information associating the physical orchemical properties of particular solvents with the production ofcrystallographic forms of drug candidates, identifying in the databasewhich physical properties are associated with producing a large numberof crystallographic forms, and designing new libraries using theseidentified physical or chemical properties as criteria for groupingsolvents.

FIG. 5 shows an illustrative flow chart of library design process 500 inaccordance with the principles of the present invention. Library designprocess 500 may be used, for example, to generate library 400illustrated in FIG. 4. At step 510, a user may define or select one ormore sources using a user terminal (e.g., user interface equipment 180of FIG. 1). Sources may include materials such as salt reactants,solvents, drug candidates, mixtures of solvents, etc. that are used toprepare a library. Also at step 510, a user may define or select alibrary layout. A library layout may represent the layout in which alibrary will be created on a substrate (e.g., a substrate with an 12 by8 matrix array of wells) or a reactor assembly. Alternatively, a librarylayout is not necessarily confined to actual physical parameters, ratherit can be provided in an intangible format (e.g., on a computer).

The user can identify source materials and library layouts by enteringidentifying information manually or by selecting identifying informationfrom a pre-defined source such as database 114 of FIG. 1.

At step 520, process 500 may define the composition of each librarymember based on the sources selected in step 510. The composition ofeach library member is defined by a mapping sequence that assignsmaterials to each library member. The mapping sequence may beautomatically generated or defined by a user. Automatically generatedmapping sequences may provide an exhaustive mapping of materialcompositions than can be used to create libraries.

When step 520 has completed defining the composition of each librarymember, process 500 optionally proceeds to step 530. At step 530, arecipe file is generated based on the composition defined at step 520.The recipe file may embody handling instructions that can enableinstruments such as material handling apparatus 150 (FIG. 1) to preparethe library. Step 530 is optional because the library mapping parameterscan be integrated with software that controls a material dispensingapparatus to prepare members of substrate according to the library.

Persons skilled in the art will appreciate that the above discussionwith respect to FIGS. 4 and 5 is not intended to be an exhaustivedescription of library designing. The discussion does, however, discussa portion of the various features pertinent to describing the presentinvention. For example, WO 00/23921, provides a substantial descriptionof library designing.

After a library is designed the process proceeds to prepare one or morelibraries using material handling apparatuses. FIG. 6 shows anillustrative flow diagram of combinatorial library preparation process600 that controls hardware to automatically prepare libraries inaccordance with the principles of the present invention. Automated stepmay be performed manually if desired.

At step 610, the user may define hardware resources that are availablefor use in process 600. The user may define hardware resources such asautomated liquid handling robots, pump controllers, solid or powderdispensing systems, and other dispensing equipment. In defining thehardware, the operating characteristics of such hardware may also beidentified. For example, characteristics such as configuration ofsyringes attached to a dispensing device (e.g., in series of parallel),motion limits, step size and reference positions for arm movements,dispensing capacity of syringes, etc. may be identified. Other hardwareresources may include temperature controllers for regulating thetemperature or temperature ramp rate of a substrate or pressure of areaction vessel.

At step 620, a recipe file or other instructions that include thelibrary design is received by process 600. Step 620 may analyze the dataand develop a set of material handling instructions that can control thehardware to synthesize a library. The material handling instructions maybe provided to, for example, electronic circuitry 112 of FIG. 1 so thatcontrol commands can be provided to material handling apparatuses. Thenat step 630, the material handling instructions are executed and alibrary is prepared in accordance with the recipe file received at step610.

Software that can perform process 600 is described in more detail in WO00/67086, which is hereby incorporated by reference in its entirety.

After salts, crystals, or polymorphs are created in a library format,they are screened for desired properties and characterization. Thepresent invention utilizes high throughput screening that providessufficient information that enables the process to proceed to the nextstep. Screening tests are performed to obtain data on each librarymember. Screening tests can be primary or secondary depending where inthe process the test are performed. A minimum number of screening testsmay be performed to identify the materials in the library, determine thecrystallinity of the materials, and determine the solubility of thematerials. Additional tests may be performed to obtain more data on oneor more library members.

Screening tests may provide quantitative and/or qualitative data.Quantitative data enables the present invention to perform analysisbased on numerical data. This data is well suited for use in softwareprograms operating on computers because it can be categorized quicklyand accurately. Qualitative data, however, provides information thatdoes not require intensive number manipulation and relatively quickmaterial characterization. For example, an identity screen can run withsufficient precision to determine if members of the library aredifferent from each other or from a standard. But the identity screenmay not be so thorough that it identifies each member of the library. Inanother example, a crystallinity screen may determine certain crystalcharacteristics, such as morphology, without determining every (or evenmost) characteristics of a crystal (e.g., such as melting point, unitcell, etc.).

Screening tests may be implemented on instruments that determine, forexample, solubility, partition coefficient (log P), birefringence (whilethe sample is wet and/or dry), melting point, crystal morphology,hygroscopicity, and other physical characteristics. Other screening testinstruments may provided data using, for example, X-ray diffraction,Raman spectroscopy, IR or Near IR spectroscopy, UV-Vis spectroscopy,nuclear magnetic resonance spectroscopy (NMR), gas chromatography andliquid chromatography. These tests are preferably performed in parallelor in a rapid serial or automated mode such that the screening methoddoes not delay the overall process.

As illustrated in FIGS. 2A, 2B and 3, screening test are performed indifferent steps of the process. One or more screening test may beperformed at each screening step in a process. Several screening stepsprovide greater quantities of data so that library members are morefully characterized than characterizations performed by a singlescreening test. For example, in one embodiment a primary screening step(e.g., step 225 of FIG. 2A) may perform at least 4 different tests(e.g., solubility, log P, crystallinity and Raman spectroscopy) todetermine which library members should be selected for bulk synthesis.Primary screening steps preferably use fewer tests than a secondaryscreening step so that high throughput screening can be maximized.

A secondary screening step (e.g., step 255 of FIG. 2B) may also performseveral high throughput screening test to provide the process with athorough characterization and identification of the bulk sample.Secondary screening test may include test such as IR spectroscopy, NearIR spectroscopy, UV-Vis absorption, X-ray diffraction, melting point,and pK_(a). Other tests that can be performed on bulk samples includeNMR, differential scanning calorimetry, thermogravimetric analysis andelemental analysis.

A process may perform solubility, birefringence X-ray diffraction,hygroscopicity and/or Raman spectroscopy in high throughput mode toquickly and accurately characterize library members. In particular, whenusing only solubility, birefringence, and Raman tests, these testsadequately characterize any salts that may have formed. In addition,these test also determine whether polymorphs of drug candidates (orsalts thereof) have formed. If desired, a more detailed identificationof the different polymorphs can then be accomplished through additionalscreening test and analysis.

Solubility is performed by sampling the supernatant mother liquor fromthe recrystallization step of polymorph generation or from thecrystallization or precipitation step of salt selection, as describedabove. The liquid sample is subjected to a concentration detector todetermine the amount of the drug candidate (or salt) in the solvent. Thedrug candidate (or salt) concentration may be detected using liquidchromatography, thin layer chromatography, gas chromatography,absorption in the UV-Vis range, infrared (IR), fluorescence or any othersuitable technique. In one embodiment, liquid chromatography coupledwith an ultraviolet radiation detector may be used to determine drugcandidate concentration. An example of an liquid chromatography systemis an Agilent 1100LC system.

Sampling for the solubility test is typically performed at a temperaturein which recrystallization or precipitation has occurred. Solubilitytesting can be performed as a high throughput screen, which can beperformed in a rapid serial mode or in parallel. The testing willprovide solubility information for the particular solvent in which thedrug candidate salt is present at which the solution was sampled. In oneembodiment, one may determine the solubility of one or more drugcandidates under a variety of conditions in parallel microtiter platesor other assembly comprising a plurality of wells by forming arrays ofsolvents or solvent mixtures at temperatures ranging from 0° C. to 70°C. and measuring the concentration of the compound in the supernatant.In a preferred embodiment, the temperature is measured at a temperaturein which the drug candidate or salt is dissolved (T_(initial)) and atthe final temperature, in which the drug candidate or salt hascrystallized (T_(final)).

The partition coefficient, also called log P, is a well known measure ofsolubility in a water/1-octanol mixture. In a high throughput mode, logP is determined by measuring the concentration of the drug candidatesalt in both water and 1-octanol at a particular temperature. In apreferred embodiment, the solubility in 1-octanol and the solubility inwater is measured in separate wells of a microtiter plate as part of theoveral process, with those measurements used to determine log P. In analternative embodiment, water saturated with 1-octanol and 1-octanolsaturated with water may be used as solvents. The concentration may bedetected using liquid chromatography, thin layer chromatography, gaschromatography, absorption in the UV-Vis range, infrared, fluorescence,or any other technique that determines concentration known to those ofskill in the art. Using any suitable detection device, log P isdetermined by dividing the concentration in 1-octanol by theconcentration in water and taking the log of that number.

Birefringence testing may be used to determine the crystallinity of asample. In particular, birefringence testing indicates the quantity ofcrystals formed, size of the crystals, and shape of crystal (e.g.,needle structure, blade structure, tabular structure, or any otherstructure). FIG. 7 shows several crystal structures that can be detectedusing birefringence testing in accordance with the principles of thepresent invention. The crystal's structure may provide information indeciding which library members are suitable for large scale synthesis.For example, needle crystals are often more difficult to filter thantabular crystals.

Birefringence testing may be performed by passing light through wet ordry samples in the library. Preferably, the samples are arrayed on atransparent substrate located between two parallel and perpendicularlyaligned cross-polarized filters. A light system may be positioned aboveor below one of the filters to detect light refraction as it passesthrough samples on the substrate. Persons skilled in the art willappreciate that other devices can be used to detect birefringence ofmaterials in such a setup. For example, an array of photo-diodes may byused to detect birefringence of a material. The light systemadvantageously screen the entire substrate in one pass, thus providing arelative fast indication of the suitability of the materials located onthe substrate. A detailed description of a birefringence testing deviceis described in detail in conjunction with FIG. 28. Also described inconjunction with FIG. 28 is a method for performing light scattering todetect if any crystalline structures are present.

Because crystals are birefringent structures, they have the ability torefract light. This property allows a birefringence testing station todetect if a sample on the substrate is crystalline or amorphous.Amorphous materials are typically undesirable because they tend to beunstable and more hygroscopic than crystalline forms. An example ofdevice using a parallel light rotating and collection device isdescribed in U.S. Pat. No. 6,157,449 ('449 patent), which isincorporated herein by reference in its entirety. A birefringencetesting station may, for example, use the device in the '449 patent inconjunction with a light system.

In situ measurements of an array of material samples can be performedusing birefringence or a light scattering technique. More particularly,in situ measurements are performed using a reflective optical scanningtechnique. A description of an apparatus that performs in-situmonitoring using the reflective scanning technique is described below inspecification pertaining to FIG. 30.

One advantage of in-situ measuring is that it maximizes high throughputtesting of material samples. For example, assume that an array ofmaterial samples are subjected to crystallization conditions while beingmonitored in situ. Further assume that these particular crystallizationcondition did not yield any crystalline structures. Instead ofdisassembling a crystallization assembly and providing the substrate,which does not contain any crystalline structure, to a series ofscreening test, in situ monitoring can provide information to avoid suchan unnecessary step. Performing screening test on material samples thatdo not contain crystalline structures slows down the screening processfor detecting new polymorphs. Thus, in situ monitoring providesinformation that may result in subjecting the material samples todifferent crystallization conditions that may produce crystals.

Hygroscopicity testing characterizes materials (e.g., crystals)according to their ability to adsorb water. Hygroscopicity can be testedin an automated manner using Raman spectroscopy, Near IR spectroscopy orin situ measurements as a series of “snapshots” to determine water gain(or loss), as described in other parts of this specification. PuuMan Oy(Kuopio, Finland) manufactures and markets the HMA (HygroscopicityMeasurement Apparatus) that is capable of measuring the hygroscopicityof eight samples simultaneously.

Hygroscopicity can also be measured by automated weighing systems. Forexample, vials containing samples are automatically weighed (for exampleusing a Bohdan Automated Weighing Station (called the BalanceAutomator), available from Bohdan Automation (a Mettler-Toledo Company,Vernon Hills, Ill.). The vials containing samples are then exposed to acontrolled atmosphere (e.g., a certain humidity) for a selected time,after which the vials containing samples are automatically weighedagain, with a difference in weight being a measure of hygroscopicity. Insome embodiments, the automated weighing station can be located inside aglove box or other atmosphere controlled chamber.

Hygroscopicity can also be measure using dielectric measurements. Suchdielectric measurements can be performed on a substrate having regionscomprising interdigitized probes, with the samples being placed in theregions and the substrate being placed in a controlled atmospherechamber (e.g., a controlled humidity chamber). Changes in samplesdielectric properties are measured as water is gained (or lost) andhygroscopicity is determined.

In a preferred embodiment, hygroscopicity is measured in a highthroughput manner using a microbalance and more particularly usingsensitive mechanical resonators, whose resonance performance can bemonitored and correlated with mass. In one preferred embodiment,hygroscopicity is measured using a method for screening samples createdin accord with the description herein (e.g., on a universal substrate),comprising the steps of (a) providing a plurality of solid samples; (b)placing a first sample onto a mechanical resonator in signaling (e.g.,electrical, magnetic, optical, thermal, or other communication)communication with a source of an input signal; (c) coupling themechanical resonator with measurement hardware; (d) exposing the samplesto a controlled atmosphere (e.g., moisture or desiccating) while on themechanical resonator; (e) applying an input signal; (f) monitoring aresponse of the mechanical resonator to the moisture of the samplesthereon with the measurement hardware; and (g) repeating steps (b)through (f) for each sample for which measurement is desired. Inaddition, this method can readily be adapted for also conductinganalysis of mass change in response to a change of temperature, such asfor thermogravimetric analysis.

In this preferred method, the monitoring that occurs in step (d) mayemploy a suitable lock-in amplifier or like hardware for monitoring thechange of frequency of the mechanical resonator while maintaining theinput signal to the resonator as a constant. It may alternatively employthe monitoring of the change in electrical feedback from the resonatorwhile maintaining a constant frequency.

In a particularly preferred embodiment, the input signal is a variablefrequency input signal and the monitoring step (d) includes varying thefrequency of a variable frequency input signal over a predeterminedfrequency range to obtain a frequency-dependent resonator response ofthe mechanical resonator. The preferred method advantageously allowsrepeating steps to be performed simultaneously for analyzing an array ofsamples in a parallel format. Yet, as desired the repeating steps may beperformed serially.

When employed in a salt selection or polymorph workflow, as describedherein, the preferred method can be described as comprising the steps of(a) providing an array of different particulated pharmaceuticalpolymorph candidate samples; (b) providing a tuning fork resonatorhaving at least two tines with tips and being in electricalcommunication with a source of an input signal; (c) adhering a quantityof a plurality of samples to at least one of the tines; (d) coupling thetuning fork resonator with measurement hardware; (e) simultaneously, forat least two samples of the array, humidifying the samples while on thetuning fork resonator; (f) simultaneously, for at least two samples ofthe array, applying a variable frequency input signal; (g)simultaneously, for at least two samples of the array, varying thefrequency of a variable frequency input signal over a predeterminedfrequency range to obtain a frequency-dependent resonator response ofthe mechanical resonator to the humidification of the samples; and (h)graphically displaying the responses for each of the samples analyzed,such as by providing a readout of a frequency response, whereinfrequency versus signal is plotted.

Another embodiment of the preferred method contemplates an apparatus formeasuring small quantities of materials, comprising a plurality ofresonators, and particularly tuning fork resonators having tines withtips; a holder for each resonator; a readout board; a plurality ofelongated members for bridging electrical communication between theresonator and the readout board; and a frame carrying at least theresonators, holders and elongated members. The apparatus is preferablyadapted for attachment to a robot arm for facilitating automation of theoperation of the apparatus. The apparatus of may further comprise othercomponents, such as a sample work surface having a recess therein forreceiving a sample, a host computer, and a power source (e.g., forproviding a variable frequency input signal to the resonators).

The advantages of the preferred method are numerous, including massmeasurements of soft, thick, non-uniform layers or irregularly shapedsamples; small sample quantity measurements (with some samples beingless than about 100 micrograms and more preferably less than about 50micrograms); certain resonators (e.g., tuning fork resonators) have aQ-factor does not decrease by more than about 1-3%, so relative changeof a sample mass is accurately measured by resonator frequency change;quick measurements, in some embodiments in less than one minute (and inother embodiments in less than about 30 seconds or less than about 5seconds for a single sample or for an entire array or library); and theability for real-time mass tracking (or real-time hygroscopicity). Thedetails of this preferred method are set forth in U.S. Pat. No.6,928,877, titled “High Throughput Microbalance and Methods Using Same”,which is incorporated herein by reference for all purposes. See alsoU.S. Pat. Nos. 6,336,353 and 6,182,499, which are both incorporatedherein by reference for all purposes.

Raman spectroscopy may be performed using any type of unit, such as acommercially availably unit (e.g., Renishaw, Ramascope), with an X-Ystage that addresses the samples in a rapid serial mode. In someembodiments, in order to run a very high throughput screen, peakassignments may not be performed on the spectra acquired. Instead, thespectra are used as “fingerprints” to determine if different polymorphs(or salts) have formed in the high throughput experimentation. In otherembodiments, peak-matching software may be used to make thedetermination of different identity. Raman, IR, X-ray or otherfingerprint type spectra may not be quantitatively analyzed, and insteadmay be used for qualitative determinations about the relative samenessor differences between spectra. An example of spectra provided in agraph format is illustrated in FIG. 9.

Morphology or crystallinity may also be performed by inspecting each ofthe regions of the libraries under a microscope, for example, withcrossed polarizers. X-ray diffraction can be performed on a Bruker GADDS(Bruker AXS, Madison, Wis.), See also, U.S. Pat. No. 6,371,640, whichdiscloses a method and apparatus of screening materials in a highthroughput and library format, incorporated herein by reference.

Thus, selected screening test are used to select drug candidates forfurther investigation. Screening test also enable the process to performdetailed characterization of library members (e.g., bulk samples) todetermine its suitability. The recrystallization condition identifiedbased on the selected samples may be used to prepare bulk samples of thedesired salts for additional characterization. One feature of theprocess of the present invention is the use of a “universal substrate”.A universal substrate refers to a substrate having samples thereon andthat can be used for a variety of tests (described above) without manualor other manipulation of the samples. This will become more apparent inthe discussion below. Thus, a single substrate (e.g., array ofmaterials) can be tested for birefringence, Raman, X-ray diffraction andmelting point without handling the samples for each test.

Automated control of material screening device 160 (e.g., birefringencestation, Raman station, XRD station, melting point station, etc.) mayadvantageously enhance high throughput screening. High throughputscreening is enhanced by automated control because it allows the processto quickly characterize drug candidates and provide data to a computer(e.g., computer 110 of FIG. 1). Automated control may be provided by asoftware program operating on a computer. In particular, softwareprograms may control material screening device 160 to characterize andidentify properties of drug candidates.

For example, a software program can instruct material screening device160 to perform an identity screen of the drug candidates. Because drugcandidates are typically arranged in a library format, the software candirect device 160 to perform screening in parallel or in a rapid serialmode. Parallel screening provides characterization of two or more drugcandidates simultaneously. Rapid serial mode screening providesrelatively rapid screening of drug candidates on an individual basis.

Data is obtained from material screening device 160 when itcharacterizes and identifies properties of drug candidates. This datamay be provided to the software so that analysis can be performed withrespect to each drug candidate. Data analysis can include categorizingdrug candidates, determining suitable drug candidates (e.g., for saltselection), experimental data categorization, and determining polymorphs(e.g., for drug candidates, salts, and other solutions). Data analysismay be performed one or more times during a process. For example, dataanalysis may be performed after an initial screening to determine whichlibrary compositions are suitable for further testing. (This isillustrated in FIGS. 2A and 3 at steps 230 and 335, respectively.) If atleast one of the library compositions is suitable for further testing,secondary data analysis may be performed in the process. Secondary dataanalysis may yield a substantially more rigorous examination of datathan primary data analysis and provide accurate results in highthroughput. (Secondary data analysis in a process is illustrated inFIGS. 2B and 3 at steps 260 and 360, respectively.) Persons skilled inthe art will appreciate that data analysis can be performed as oftentimes as necessary to characterize and examine data.

FIG. 8 shows an illustrative flow diagram for determining if acrystalline structure being tested is part of a particular polymorphfamily or is a newly discovered polymorph. Categorization flow 800determines if any material compositions in the library member should beaffiliated with an existing family of materials (e.g., existingpolymorph family) or if it should be placed in a new family of materials(e.g., new polymorph family). A family comprises a group of materials(e.g., crystalline structures) that exhibit similar characteristics. Thecategorization of various library members is performed based on dataobtained from screening tests (e.g., Raman, XRD, melting point,solubility, hygroscopicity, etc.). One advantage of analysis flow 800 isthat it can categorize library members (e.g., hundreds or thousands ofmaterials) into an appropriate family (i.e., existing or new) withoutrelying on initial reference data. Initial reference data can also beused, and in some embodiments, reference data is preferred, such as whena known polymorph exists and others are being searched. Categorizationflow 800 builds a database of reference data (e.g., existing polymorphfamilies) as it categorizes library members with the database beingintegrated with known or existing data or not. Based on this database,categorization flow 800 continuously analyzes and categorizescrystalline structures created using, for example, process 200 and/orprocess 300, as described herein.

In the following discussion of FIG. 8, assume that there is no priordata for any of the crystalline structures discussed in this particularexample. At step 810, data obtained from a screening test of crystallinestructures of the library array is provided. Any above-describedscreening test can be used to provide data on each crystallinestructure. It is preferred that the data set is an XY dataset. Forexample, if Raman, XRD or another spectroscopic screening technique isused, spectra is provided. Spectra provides data such as peak location,peak height, and peak width of a crystalline structure. If, for example,a melting point screening technique is used, numerical temperaturevalues are provided as data to step 810.

Assuming that the data obtained at step 810 is the first set of dataobtained for the first crystalline structure, there is no reference datathat can be used for comparison. Thus database at step 820 is void ofreference data, at least initially. In particular, step 820 does nothave any reference data that represents a polymorph family. For purposesof clarity and brevity, any data that is provided from step 810 that isused in process 800 is referred to as new crystalline structure.Proceeding to step 830, the data associated with a crystalline structureis compared to data for each known polymorph family stored at step 820.Thus, the crystalline structure undergoes an iterative comparisonprocess, which after each iteration, produces a correlation coefficient.A correlation coefficient is indicative of how “close” the crystallinestructure is to a particular polymorph family.

Persons skilled in the art will appreciate that methods other thaniterative techniques can be implemented to compare the crystallinestructure to each of the existing polymorph families.

Categorization process 800 may use statistical methods to obtain thecorrelation coefficient. Statistical methods may be used to determinethe deviation (e.g., standard deviation) of the data associated with thecrystalline structure to a reference data set. Software programs thatperform such mathematical functions include MATLAB® software sold by TheMathWorks, Inc., of Natick, Mass., Mathemathica® sold by WolfrumResearch, Inc. of Champaign, Ill., and MathCad® sold by MathSoft ofCambridge, Mass. Persons skilled in the art will appreciate that othersoftware programs different from the programs described above may beused to perform matrix based calculations and other mathematicalcalculations.

The comparison process in step 830 uses different comparison techniquesdifferently based on the type of data provided by step 810. One suchtechnique is a cross-correlation technique. This technique is typicallyused when the measured data is obtained using Raman spectroscopy. Asthose of skill in the art know, Raman spectra it typically a curve orgraph that represents characteristics of a crystal. Such a graph isillustrated in FIG. 9. The cross-correlation technique performs apoint-to-point correlation to determine how “close” the measured spectrais to reference spectra. After the comparison is performed, acorrelation coefficient is obtained based on how close the graphs match.

Another technique that can be implemented at step 820 is a peak matchingpercentage technique (e.g., peak locations are compared). This techniqueis typically used comparing spectra obtained by X-ray diffraction (XRD)spectroscopy. In this technique, the peak locations of an XRD graph aredetermined using an algorithm. Then the determined peak locations arecompared to peak locations of reference data (e.g., peaks of particularpolymorph family). A percentage value is obtained based on how similarthe XRD peaks are to the reference peaks. This percentage value isanalogous to the correlation coefficient.

After the iterative comparisons are performed, the best correlationcoefficient is obtained at step 840. The best correlation coefficient isassociated with the polymorph family that the new crystalline structureset matched best.

At step 850, the best correlation coefficient is compared to apredetermined value. Typically, the predetermined value is auser-defined correlation coefficient that sets the threshold fordetermining whether the data associated with the crystalline structureshould be associated with an existing polymorph family.

If the correlation coefficient exceeds the predetermined value, thatcrystalline structure is grouped into the polymorph family associatedwith that best correlation coefficient at step 860. After step 860,process 800 returns to step 810, which provides the next new crystallinestructure to step 830.

If the correlation coefficient does not exceed the predetermined value,a new crystalline polymorph family is created based on the data obtainedon the new crystalline structure at step 870. In the event that therewas no reference polymorph family for any comparison to be performed,the crystalline structure is automatically used to create a newpolymorph family at step 870. The data associated with the new polymorphfamily is provided to step 820 for use as a reference as an existingpolymorph family. Also, after step 870, process 800 returns to step 810.Data from a whole series of libraries based on a single drug candidatecan be sorted in continuous process so that one set of families arecreated that are indicative of the individual forms of the drugcandidate.

Thus, analysis process 800 illustrates crystal structure categorizationbased on screened data. Persons skilled in the art will appreciate thatsteps shown in FIG. 8 are merely exemplary and that additional steps maybe added and some steps may be omitted or modified.

If desired, reference data such as a computer file, a look up table, orother suitable information source may be provided to the database instep 820. Parameters that were used to create the library members can beprovided to database in step 820. For example, parameters such assolvents added to the salt, crystallization temperature, and otherparameter used to formulate library arrays and crystallize the arrays.This data can be used as criteria in grouping crystalline structures.

Storing measured data provides a database that stores data (e.g.,spectra) from each library member. This advantageously enables currentlymeasured data to be compared to other previously screened libraries. Insome embodiments, when a new family of polymorphs is discovered in step825, data from previously identified families may be compared to newlyacquired data. This enables analysis process 800 to determine if the“new” family corresponds to a previously identified family such that thedata can be assigned to an existing family. This effectively reduces thetotal number of families of data and allows for a high correlationwithin a family or group of data. Yet, user defined variation betweenfamilies or groups is still preserved. In other embodiments, if originaldata is not provided, the first experimentally determined data can beused as the “starting” information (e.g., step 820 can use data from adesignated well or a designated piece of information).

An illustrative example of pre-formulation system 100 (FIG. 1) thatimplements process 800 to categorize different library members isdescribed in conjunction with FIGS. 9-12. In the following example,assume that each library member of a 96 well substrate has been testedusing a device (e.g., an infra-red device, UV-Vis absorption device, aRaman device, an X-ray device) that obtains data. Using such devices,the data can be arranged in a graphical format. FIG. 9 illustratesspectral data obtained from four different library members using a Ramandevice. The spectral data shows that each of the library members containpolymorphs of the same drug candidate, but are similar enough to beclassified as part of the same family or group.

FIG. 10 illustrates spectral data obtained from three different librarymembers using, for example, an X-ray diffraction device. FIG. 10 showsthat these three library members also have polymorphs, but are eachdifferent from each other such that they are not part of the same familygroup.

FIG. 11 shows an illustrative interactive display screen 1100 thatincludes several spectral graphs of screened library members. Displayscreen 1100 shows that spectra 1105 are arranged in array format 1108.Preferably, spectra 1105 are arranged so that they coincide with thelayout of the substrate in which they were screened. As shown in FIG.11, not every portion of array 1108 has spectral element. This may bebecause a crystal did not form during the crystallization (e.g.,recrystallization) process.

FIG. 11 also shows that a user can enter and change spectrum sortingparameters 1110. For example, a user can enter a minimum groupingcorrelation coefficient. As described above in conjunction with FIG. 8,the correlation coefficient may be used to determine if a crystallinestructure belongs to a particular family. The value entered for thecorrelation coefficient can range between −1 and 1. The closer thecorrelation coefficient is to 1, the more stringent the criteria becomesfor placing a particular library element in an existing family. Forexample, if there are ten library elements and the correlationcoefficient is 0.9, each library element may be associated with a newfamily—thus creating ten different families. If the correlationcoefficient is closer to zero, then matching a particular libraryelement to an existing family becomes less stringent, thereby producingfewer new families. The value selected for a correlation coefficienttypically ranges from about 0.5 to about 0.9. The correlationcoefficient selected may, for example, be the predetermined value usedat step 850 in process 800.

A user can select whether to use a fixed reference (e.g., apredetermined reference) or an arbitrary reference (e.g., a libraryelement) for providing a baseline in determining family groupingselection. Persons skilled in the art will appreciate that additionalparameters may be entered or modified as suitable within the spirit ofthe invention. A user may also change the image size of array 1108 bychanging width and height parameters 1115. Users can submit theirentries by selecting send overlay 1120 or they can reset their entriesby selecting reset overlay 1122.

In another embodiment, categorization process 800 may be implementedwith an XY dataset where similarity among xy data sets is measured by acorrelation coefficient (CC) ranging from −1.0 to +1.0, with +1.0 beinga perfect match to be in the same group, which is used for data deemedof insufficient quality. The sorter has a default group, namely the junkgroup. The parameters for determining the data to be junk or not can beset by the user, with each data set being checked to see if it is junkbefore being placed in this default location. When a data set is not putinto the default location, a new group may be created by comparing thexy data set from the remaining data to a reference and for eachunclassified sample xy data set using the reference set from eachexisting group (or one can pull out all sets from each existing group).Comparison of the sample data to the reference set can provide a CCvalue (or comparison of the sample data to the sets and get a best CCvalue from the comparisons). This is followed by retention of the bestCC and the Group where the CC comparison came from. If the best CC isless than the specified value, then the sample is put in the new group;otherwise the sample is put into the group from which the best CC wasobtained. The user can override the automated classification byperforming comparisons visually and assigning groups manually. This canalso be implemented on peak locations, peak heights or other data. Thus,for XRD data, it is currently preferred that peak locations are used todetermine a correlation coefficient.

In another embodiment, the process also allows the user to define arecycle bin, where data can be designated for re-running through thecorrelation process. In this manner, the user can identify data thatshould be sorted based on different parameters. In still anotherembodiment, the user can iterate the correlation process, by runningmultiple correlations based on at least one different parameter.

FIG. 11A shows one example of a dialog box 1150 for the user to definethe parameters used in correlating the spectra. One side of the box 1150allows the user to define the automatic junk classification 1152, whichas described above allows for removal of data from the classificationworkflow. The do it check 1154 enables or disables this feature, withthe settings that the user can define included in the remainder of thebox 1152. The moving average point 1156 is used for smoothing the signalof the data. The minimum peak height 1158 is used for setting the heightthat a peak in the xy data set must meet in order to be considered to bea peak for correlation purposes (e.g., this feature can set backgroundnoise levels and allow the correlation process to ignore the noise). Theminimum zero crossings 1160 allows the user to set the minimum number ofzero crossings for peaks, with the implicit assumption that each peakhas two zero crossings as a default. The maximum zero crossings 1162allows the user to set the maximum number of zero crossings in a dataset so that the data would be considered to have good quality. Forexample, spectra with too many zero crossings may signify that thespectra may simply comprise noise.

The match algorithm 1164 side of box 1150 is used for setting thecorrelation factors. The coverage 1166 has settings for single and all,with single comparing the sample to a master or reference data of theexisting family or form only. The resulting score represents thecomparison score between the sample and the reference data for aparticular family. The all setting compares the sample data to all ofthe data already in the family, in which case that best correlationscore represents the best score resulting from all the comparisons. Themethod 1168 allows the user to decide on what part of the data will beused for correlation. As shown in FIG. 11A, peak center performscorrelation based on the peak locations (within an acceptable deviationas defined in the peak center band width 1172). Other methods ofcorrelation include full signal correlation, which uses the entire dataset for correlation. Differential signal method of correlation performscorrelations based on the digital difference signals (first derivative)of the sample and reference data. Baseline removed method of correlationperforms correlation based on data with the curvatures and slopesremoved in the data. Other methods of correlation are within the skillof one of skill in the art, including for example with spectra, peakheight or peak width. The typical correlation coefficient 1170 is thesetting to determine the allowable difference to obtain the bestcorrelation coefficient. The peak center band width 1172 allows for theuser to set an allowable deviation in peak centers in spectra data sets.The minimum peak height 1174 generally performs the same function as1158, and moving points average 1176 allows the user to average setnumber of data points together for correlation purposes.

FIG. 11B shows an example of a dialog box 1180 that allows the user toset the parameters for performing the correlation. The user sets theminimum correlation coefficient, as discussed above, with box 1182. Theuser can allow for previous sorting results to be used in a new sort byeither checking or un-checking box 1184, with use of previous sortresults allowing for re-cycling the data, as discussed above. The regionof interest 1186 portion allows the user to define a certain portion ofthe data to consider in the correlation workflow, for example if only acertain region of a spectra would be of interest, then this portion canbe used for correlation. As shown in box 1180, this particularembodiment allows the user to set up to five interval ranges 1190 (witha lower setting (low X) and an upper setting (high X)).

FIG. 11C shows dialog box 1192 that allows the user to manually movedata from one group (e.g., family or form) to another group (or family).This feature allows the user to manually create families used forcorrelation or to manually change correlations.

When the user selects send overlay 1120 (FIG. 11), the process mayanalyze the spectral data in accordance with the parameters defined bythe user. For example, the process may utilize the principlesillustrated in FIG. 8 to analyze and group each library element to anexisting or a new family. FIG. 12 shows an illustrative display screen1200 that has organized each of the library elements of FIG. 12according to their respective family. In particular, display screen 1200shows that library elements are grouped into twelve different families.Thus, each of the library elements are grouped accordingly.

A user may repeatedly change the correlation coefficient and select sendoverlay 1120 to generate different groupings of library members inspectra 1105. For example, if user desires to generate six families oflibrary elements, the user may change the correlation coefficientachieve such a result. A user may desire to do this because a particularnumber of families is known or expected. The user may know how manyfamilies are expected because a different test (e.g., a Raman,hygroscopicity, XRD, melting point) previously yielded such a result.

One advantage of using the software described herein is that a user canrun several categorization tests based on different data sets. Forexample, a user can categorize library members based on data obtainedfrom, solubility, log P, crystallinity, melting point, hygroscopicity,crystal morphology and birefringence, as well as X-RAY diffraction,infrared (IR), Near IR, and Raman spectroscopy, among other screeningtechniques. After categorization is performed based on two or more suchtests, the results of each categorization can be cross-referenced todetermine inconsistencies and/or to validate findings of potential newpolymorphs.

As described above in conjunction with FIGS. 2A, 2B, and 3, processes200 and 300 prepare and screen, and analyze library members to groupdifferent polymorphs. Because it is an object of the present inventionto provide high throughput testing of polymorphs, software can beimplemented to optimize the screening process. Software can minimize thenumber of screening test that need to be performed for a particularlibrary member. Thus, increasing the capacity of the number of libraryelements that can be tested for polymorphs.

FIG. 12A shows an illustrative flow diagram of a software process 1270that determines which library members should be subjected to furthertesting in accordance with the principles of the present invention.Process 1270 begins at step 1272 in which each of the library membersare subjected to an initial screening. Screening, as described above,provides data to a computer, and based on that data, process 1270 candetermine whether a particular member is suitable for additionaltesting. Preferably, the initial screening is a relatively fastscreening such as birefringence testing.

At step 1274, process 1270 determines whether the data associated with aparticular library element meets certain criteria. The criteria, whichis provided by step 1275, includes data that provides a basis orthreshold value for selecting which library member should be selected toperformed additional testing. For example, if birefringence testing isused, the criteria may be whether any type of crystal structure ispresent in the library member. The criteria that can be selected forsetting the threshold can be selected by a user or the computer programmaking the determination.

In one embodiment, later screening tests (such as Raman, XRD, etc.) canbe performed only on samples having a birefringence image with anarithmetic mean above a determined point. The arithmetic mean may be inthis case the mean of all the pixel intensities from an image of thelibrary member under birefringence conditions.

If the data shows that a crystal structure is present in the librarymember, then that member is suitable for additional testing at step1276. If, however, the data indicates that no crystal structure ispresent in that library member, that member is marked as not suitablefor additional testing at step 1278. Once a library member is selected,more screening tests (e.g., Raman, XRD, melting point) may be performed.Because performing additional tests take time, process 1270 efficientlyeliminates unnecessary testing, thereby maximizing the throughput of theoverall process (e.g., workflow).

If desired, the selective screening process can be progressive. That is,assume that a particular library element was selected in an initialscreening and was subjected to a second screening (e.g., an XRDscreening). Applying the principles described in process 1270, the samelibrary member does not have to be subjected to additional screeningtest if it fails to meet a minimum criteria standard based on the secondscreening.

The following describes several apparatuses that are used to implementprocesses for selecting salts and discovering polymorphs. In addition,the following discussion describes how such apparatuses are used inconjunction with the processes illustrated in FIGS. 2A, 2B, and 3.

Processing of the selected drug candidate or salt(s) for discovery andcharacterization of suitable forms requires at least two, but preferablythree steps, to be carried out in a combinatorial or high throughputmode. The two required steps are dissolution of the drug candidate orsalt(s) thereof, and crystallization of the candidate from solution. Theoptional third step that can occur between the dissolution andcrystallization steps is separation of any remaining solids from thesolution by filtration or centrifugation. This optional step may benecessary to eliminate nucleation sites for the crystallization step.

A general reactor assembly is shown in FIG. 13. A reactor assembly isused for formulating drug candidates, solvents, acids, bases, etc.Reactor assemblies can be used to dissolve solutions for salt selectionor to dissolve solutions for polymorph testing. Reactor assemblies suchas reactor assembly 1300 of FIG. 13 are suitable for containingreactions of interest.

FIG. 13, for example, illustrates a cross-sectional view of reactorassembly 1300 that can be used for preparing library members inaccordance with the principles of the present invention. As illustratedin FIG. 13, reactor assembly 1300 is constructed in microtiter format.Reactor assembly 1300 includes reactor block 1302 that is constructedwith one or more wells 1304. Each reactor block 1302 can receivereaction vessels 1306. Materials such as drug candidates and solventsare mixed in reaction vessels 1306. Reaction vessels 1306 are isolatedfrom one another to prevent cross-contamination among the reactionvessels. This can be accomplished by securing sealing sheet 1308 overreaction vessels 1306 by fastening cover plate 1310 to reactor block1302 with fastening device 1312 (e.g., a bolt, screw, clamp, etc).

Persons skilled in the art will appreciate that any suitable number ofwells 1304 may be constructed. For example, a reactor assembly may have96 wells or it may have 384 wells.

Reactor block 1302 and cover plate 1310 may be constructed of anysuitable material such as metals (e.g., steel, aluminum), plastics, andceramics. Materials such as aluminum or an aluminum alloy may bepreferred because they have desirable thermal and structural properties.Reaction vessels 1306 may be plastic or glass, with glass beingpreferred. Sealing sheet 1308 is typically made from a material that ischemically resistant to the reaction of interest taking place inreaction vessels 1306 as well as being elastic for its sealingproperties. Sealing sheet 1308 may be constructed from materials such asTeflon®, silicone rubber, Vitron®, Kalrez®, or equivalents. If desired,sealing sheet 1308 may be constructed with two or more such materials.

Materials can be dispensed into reaction vessels 1306 by hand or byautomated robots. Automated equipment may increase the speed andaccuracy of step 212 (FIG. 2A). Liquid handling robots such as thosesold by Cavro Scientific Instruments, Inc. of Sunnyvale, Calif. may beused for automatically dispensing materials. The description associatedwith FIG. 6 explains in more detail how the present invention cancontrol robots to dispense materials into reaction vessels 1306.

FIG. 13 provides a general example of one such reactor assembly that canbe used in the present invention. FIG. 15 illustrates a more detailedreactor assembly that can be used for formulating a library. FIG. 15 isdiscussed below in the apparatus section of the detailed description.Also described in the Apparatus section are other assemblies and toolsthat facilitate preparation of materials. For example, FIGS. 24-28illustrate heating devices that heat materials contained within thereactor vessels.

Heating and/or agitation of the reaction mixtures in each vessels isoften used to promote dissolution of the candidate material in thesolvent. Prior to dissolution, mixing/stirring balls or magneticstirrers (e.g., fleas) may be added to the reactor by hand or with adevice such as that shown in FIG. 14 (as discussed in detail below). Thereactor block may then be placed on a rocking, rotating, or vortexingplate that is fixed with a heating element for mixing and heatingreaction contents. The magnetic stirring apparatus described above mayalso be used to agitate the materials. Also, the heating element may beprogrammable to provide a desired heat cycle.

Adding a glass or metal ball to each vial can improve agitation duringthe dissolution/synthesis step. FIG. 14 illustrates both athree-dimensional view and an exploded three-dimensional view of objectdispensing assembly 1402 that dispenses several objects in parallel inaccordance with the principles of the present invention. Dispensingassembly 1402 can be secured over a reactor assembly such as reactorassembly 1300 using flaps 1409 that extend from sidewalls 1408 to lineup with the outer edges of reactor block 1302 (shown in FIG. 13).

Dispensing assembly 1402 has a cover 1404 that covers a cavity 1406formed by sidewalls 1408 and bottom plate 1411. Bottom plate 1411 ispreferably constructed with a plurality of holes that align toreceptacles (e.g., wells or vials) in batch reactor 1300 when dispensingassembly is attached to batch reactor 1300. Also included in dispensingassembly is a separation assembly 1410. Separation assembly 1410includes isolation plate 1412 and sliding plate 1414. Isolation plate1412 may have an array of guide holes that guides an object (e.g., ballor magnetic stirrers) to the holes of sliding plate 1414. The operationof dispensing assembly 1402 will be more apparent in the followingparagraph.

An excess of objects (e.g., balls) is held in cavity 1406. From thisexcess of objects, a single object is placed in each guide hole ofisolation plate 1412 by shaking assembly 1402. A single object istransferred into each hole of sliding plate 1414 by moving sliding plate1414 to a first position that aligns the holes in it with the guideholes in isolation plate 1412. In this first position, the holes insliding plate 1414 do not align with the holes in bottom plate 1411. Theobjects cannot be dispensed directly into the receptacles in the reactorbase below, therefore, one ball is retained in each hole of slidingplate 1414. Only one ball is retained in sliding plate 1414 because itis constructed with a thickness that substantially equal to thethickness (e.g., diameter of ball) of the object. Sliding plate 1414 isthen moved to a second position which aligns the holes therein with thewells or vials in the reactor base below (via the holes in bottom plate1411), and simultaneously moves these same holes out of alignment withthe guide holes in isolation plate 1412. This permits the objectcontained within each hole in bottom sliding plate 1414 to drop into thecorresponding receptacle below. It should be noted that dispensingassembly 1402 can be modified to dispense different diameter balls bychanging separation assembly 1410. The parallel dispensing device shownin FIG. 14 may be used for dispensing other small solids such asstirring fleas.

Regardless of whether agitation objects (e.g., balls) are added to thereactor assembly, an array of solvents or solvent mixtures is added tovials contained within the reactor assembly. The contents are sealed intheir respective vials and then agitated at a specified temperature fora predetermined period of time. The temperature typically ranges fromabout 20° C. to about 10° C. lower than the boiling point of the mostvolatile solvent in the array. The time period typically ranges fromabout one hour to about 24 hours. In addition, the mixture may beaccelerated by mechanical agitation. In a preferred arrangement thesolution is mixed through the use of a vortexer, sonicator, shaker,incubator, or other suitable shaking and heating devices.

Once the solution is fully mixed, the supernatant liquid may be isolatedfrom any residual solid either by hot filtration or by centrifugation.The concentration of the solid dissolved in the supernatant liquid canthen be measured by using any suitable technique such as liquidchromatography, gas chromatography, thin layer chromatography, infraredor Raman spectroscopy, and UV-Vis adsorption (as discussed below).

After the materials are mixed, the cover of the reactor assembly isremoved and aliquots of the solution (typically a supernatant or motherliquor) are removed and transferred to a glass substrate or a universalsubstrate and allowed to cool. Glass substrates such as borosilicatereactor plates sold by Zinsser Analytic GmbH of Frankfurt, Germany, maybe used in the process. As the solution cools, crystals may form. Therate in which the substrate is cooled can be controlled. For example,the substrate may be subjected to a chilled bath, a thermal module, or achilling incubator which is sold by Torrey Pines Scientific, Inc.,Solana Beach, Calif. Alternatively, particularly when volatile solventsare used, evaporation techniques may be used to generate crystals. Ifdesired, precipitation and slurry techniques can also be used.

After cooling, evaporating, precipitating, or slurry, the supernatantmay be removed in a suitable manner (e.g., by aspirating with a pipetteand/or wicking away the solvent with filter paper). Supernatant removalcan be performed either manually or automatically using a computercontrolled device. Supernatant removal can also be performed in serial(e.g., rapid serial mode) or in parallel (e.g., a twelve tip pipette canbe used for parallel pipetting). The supernatant and the crystals canboth be analyzed for their composition, identity, and properties.

The above description described how a reactor assembly and substrate areused for performing the processes of the present invention. Thefollowing embodiment describes various assemblies that advantageouslyprovide generation of and high throughput screening of salts andpolymorphs. FIGS. 15, 16 and 21 illustrate three different assembliesthat have several components in common. Although three assemblies areshown, those of skill in the art will recognize that the functions ofthese assemblies may be combined into two assemblies or into oneassembly. Three assemblies are used herein to provide a process that canbe implemented relatively quickly using each assembly. When theassemblies of FIGS. 15, 16, and 21 are coupled for use with automaticrobots, the process may be performed more quickly and accurately.

FIG. 15 illustrates reaction assembly 1500 which is in accordance withthe principles of the present invention. Reaction assembly 1500 is usedfor a dissolution process, which can include, for example, mixing anarray of solvents to a drug candidate and heating the mixture to preparehot saturated solutions. In addition, dissolution processes can includemixing acidic or basic reactants to a drug candidate to form salts.Moreover, the mixture of chemicals added to formulation assembly 1500may based on a library model.

Reaction assembly 1500 is generally similar to batch reactor 1300 inFIG. 13, but is different in several respects. Reaction assembly 1500includes a bottom plate 1510 that has a shock-absorbent layer 1512positioned on top. Shock-absorbent layer 1512 may include a foam pad orother elastic material. Positioned above these parts is reactor base1514, which is constructed with an array of holes that can receive vials1516. Reactor base 1514 and shock-absorbent layer 1512 are secured tobottom plate 1510 using bolts 1511. A barrier sheet 1518 and a septumsheet 1520 are positioned over vials 1516. Barrier sheet 1518 isdesigned to seal the top openings of vials 1516 and to prevent solventsfrom mixing with other vials. Barrier sheet 1518 may be made of Teflon®or other suitable material. Both barrier sheet 1518 and septum sheet1520 may be pierced by a needle or cannula. Reactor cover 1522 is placedover septum sheet 1520 and has holes 1523 therethrough to allow a needleor cannula to pass through. Reactor cover 1522 is secured to reactorbase 1514 with bolts 1521.

Although bolts 1511 are specifically discussed herein, other means suchas clips, clamps or vices for securing parts of reaction assembly 1500can be used. FIG. 15 also shows a registration pin 1515 that allowsparts (e.g., reactor base 1514, barrier sheet 1518, etc.) to be alignedwhen being secured together. Several registration pins 1515 may be usedto provide accurate alignment.

Vials 1516 pass through the holes of the reactor base 1514 so that thebottom of vials 1516 rest on shock-absorbent layer 1512. Typically vials1516 are closed at the bottom (i.e., like a test tube with a rounded orflat bottom). Shock-absorbent layer 1512 is made of a resilient material(such as foam) that provides resistance to vertical displacement whendownward pressure is applied to vials 1516. Thus, assembly 1500 isolateseach vial when the reactor parts are secured together and providestolerance for dimensional differences among vials 1516. FIG. 15 shows 96vials in an eight by twelve array, but those of skill in the art willrecognize that a different number or arrangement of vials may be used inany particular experiment or format.

If desired, ball dispensing assembly 1402 (shown in FIG. 14) may be usedto dispense balls into each vial 1516 prior to sealing reactor cover1522.

FIG. 16 illustrates a partially exploded view of filtration assembly1600 which is in accordance with the principles of the presentinvention. After the mixture is prepared in reaction assembly 1500,aliquots from each vial may be placed in filtration assembly 1600.Filtration assembly 1600 separates particles (e.g., supernatant) of thealiquot, thereby providing a “seedless” solution of the aliquot that canbe tested. In addition, filtration assembly 1600 facilitates preparationof an array of parent solutions, which may be daughtered for additionaltesting.

Liquids are taken from individual wells from the reaction assembly 1500using a manual or automated pipetting instrument (as described herein)and transferred to the corresponding vials in the array of wells infiltration assembly 1600. The bottom portion of filtration assembly 1600includes bottom plate 1610, fasteners 1611, shock-absorbent layer 1612,reactor base 1614, and vials 1616. These parts (1610-1616) may be thesame parts (1510-1516) as that of the reaction assembly 1500. Thisallows for common parts to be prepared and used in different assemblies,which reduces manufacturing costs.

Filtration assembly 1600 differs from reaction assembly 1500 by theaddition of filter subassembly 1630 and top plate 1624. In addition,filtration assembly 1600 contains a sealing layer 1617 that is similarto barrier sheet 1618 except that sealing layer 1617 has pre-formedholes for allowing the filtered solution to enter vials 1616. Theseholes have a slightly smaller diameters than vials 1616, thus whenfilter subassembly 1630 is secured to base 1614 such that sealing layer1617 provides a seal between vials 1616.

Filter subassembly 1630 in FIG. 16 has two primary functions. Filterassembly 1630 facilitates filtering of the samples received fromreaction assembly 1500, and it allows sampling of the filtrate fromvials 1616. These functions can be accomplished by combining filtersubassembly 1630 with a top plate 1624. Top plate 1624 provides at leasttwo positions for a cannula or needle to deliver or aspirate solutionsto vials 1616 in filtration assembly 1600. This arrangement allows forfluid communication between vials 1616 and the top of filter assembly1600. Specifically, top plate 1624 has holes 1625A and 1625B thatcorrespond to some of or all of vials 1616 in reactor base 1614. Forexample, hole 1625A may be used to introduce fluid and hole 1625B may beused to withdraw fluid from vials 1616. Details of the apparatus forintroducing solutions to be filtered into vials 1616 or withdrawingfiltered solution from vials 1616 are discussed below.

Filter subassembly 1630 is shown in detail in FIG. 17. Filtersubassembly 1630 includes a bottom mid-layer 1632 to support filterlayer 1634. Filter layer 1634 is used to separate nucleation sites orother undissolved materials from the solution so that a substantiallypure solution is provided to vials 1616 (shown in FIG. 16). Filter layer1634 may be populated with an integrated filter and gasket 1635. Sealingplate 1636 is fitted over filter layer 1634 and provides support for topmid-layer 1638. Bottom mid-layer 1632 has receiver hole 1633A andadjacent hole 1633B located over the upper opening of some or all of thevials or wells in reactor base 1614 (FIG. 16).

Holes 1633A and 1633B permit introduction and withdrawal, respectively,of a solution to vials 1616 (FIG. 16). Sealing plate 1636 supports alarge o-ring 1740 and small o-ring 1744 for each region or vial. Whenfilter subassembly 1630 is assembled, each large o-ring 1740 forms aseal between sealing plate 1636 and top mid-layer 1638. This seal,formed by large o-ring 1740, prevents vapor crosstalk between vials1616. Another seal is formed when a needle passes through small o-ring1744. Persons skilled in the art will appreciate that other materialsmay be used to provide seals. For example, seals may be formed withvalves such as a Merlin valve sold by Merlin Instrument Co. of HalfmoonBay, Calif.

FIG. 18 illustrates o-ring sheets 1850 that may be positioned on sealingplate 1636 (FIG. 16) in accordance with the principles of the presentinvention. Large o-ring 1851 and small o-ring 1852 can be manufacturedin sheets for easier positioning on sealing plate 1636. As illustrated,o-ring sheet 1850 is constructed in a four-by-four array of o-rings.Thus, if a reactor base has 96 wells, then six o-ring sheets 1850 wouldbe required to fully populate a 96 well assembly. Other sizes of o-ringarrays may be used based on the size of each o-ring array and theoverall number of regions or vials in the reactor assembly. For example,the o-rings may be arranged in an 6×8 array, an 8×8 array, or any othersuitable combination.

FIG. 19 shows a close-up isometric view of a portion of FIG. 17. Inparticular, FIG. 19 shows first channel 1920 and second channel 1930that are formed when filter subassembly 1630 is assembled. First channel1920 may correspond to hole 1633A, which may be used for introducingliquid into the filtration assembly. In addition, FIG. 19 shows filter1634 positioned between bottom mid-layer 1632 and sealing plate 1636.Thus, when liquid is deposited into first channel 1920 via hole 1633A,it is filtered before being deposited in the receptacle. Second channel1930 may correspond to hole 1633B, which provides a path in which aneedle or cannula can directly withdraw filtrate.

In an alternative embodiment, filters may be cut to size so thatindividual filters are placed into bottom mid-layer 1632, instead ofsingle sheet of filter paper. Filter paper may be cut to size by knifeedge ring 1921 that are part of bottom mid-layer 1632. Knife edge ring1921 may, for example, be associated with the first channel. When a userplaces filter paper on bottom mid-layer 1632 and impresses it onto eachknife edge ring 1921 (only one shown), an array of filter disks onmid-layer 1632 is provided.

FIGS. 20A and 20B illustrate how pipette system 2060 dispenses intoand/or retrieves materials out of vials 1616 using filtration assembly1600. In particular, FIG. 20A shows a cross-sectional view of filtrationassembly 1600 and pipette system 2060 that has needle 2020 extendingthrough hole 1625A. Passing needle 2020 through hole 1625A enables theprocess to utilize filtration assembly 1600. FIG. 20B illustrates anenlarged view of filtration assembly 1600 encircled with dashed line“6B” in FIG. 20A. Here, needle 2020 is inserted into hole 1625A suchthat it pierces barrier sheet 1618, septum sheet 1620, and protrudesthrough small o-ring 1744. As described above, when needle 2020 andsmall o-ring 1744 interact, a seal is formed. This seal enables solventto be dispensed into hollow channel 2060 without risk of contaminatingother vials. Solvent is dispensed into hollow channel 2060 and is passedthrough filter layer 1634 or, alternatively, through a filter disk (notshown) to receiver 2061. Receiver 2061 is located in bottom middle layer1632 and is typically made of chemically resistant material (e.g.,plastic) that is designed to prevent wicking away of the solvent passingthrough filter layer 1634.

The seal between the needle 2020 and small o-ring 1744 should be capableof withstanding sufficient pressure to allow the liquid to be injectedinto the channel 2060. Bottom mid-layer 1632, sealing plate 1636, andtop mid-layer 1638 are typically made of metal (such as stainless steelor aluminum). The filter material in filter layer 1634 or filter diskcan be made of an appropriate filtering material that is stable withrespect to the solvent media, including materials such as glassmicrofiber filter pads, cellulose, Teflon®, paper, and other materialsused for sample filtration with organic solvents. Filter layer 1634 cancomprise a single sheet of filtering material so long as the process isrun to avoid cross talk between different array members. Alternatively,several pre-cut filters may be positioned above channel 2061.

FIG. 20C illustrates how pipette 2060 can either dispense or retrievesolvent directly from vial 1616. For example, needle 2020 can extendthrough hole 1625B, barrier sheet 1618, septum sheet 1620, and filterassembly 1630 to directly access vial 1616. This advantageously providesfluid communication between needle 2020 and vial 1616 without usingfiltering sheets of filtering assembly 1600.

Pipette system 2060 can dispense or retrieve liquids at a specifiedtemperature to prevent undesired crystal formation or precipitation.Pipette system 2060 may utilize, for example, heat sink 2042 and acartridge heater (not shown) (e.g., a FIREROD heater) to maintain aspecified temperature. A cartridge heater may be housed in cavity 2043.Extending from the needle 520 is a wire (not shown) that is wound aroundheat sink 2040. Because the wire is coupled to heat sink 2040, heatgenerated in heat sink 2040 is conducted to needle 2020. Thus, needle2020 is maintained at a temperature that prevents aspirated liquid fromcooling. Other designs may be employed to maintain the liquid at thedesired temperature, such as a heated tip on a needle, as shown in U.S.Pat. No. 6,260,407, which is incorporated herein by reference in itsentirety.

FIG. 21 illustrates crystallization assembly 2100 that is designed topermit the formation of crystals in accordance with the principles ofthe present invention. Mixtures may be obtained from vials 1616 offiltration assembly 1600 and dispensed in crystallization assembly 2100.Alternatively, mixtures may obtained from vials 1516 of reactionassembly 1500 (shown in FIG. 15). Crystal formation may occur, forexample, by cooling the mixture, evaporating the mixture, byprecipitating the mixture, or by slurrying the mixture.

One benefit of having separate filtering and crystallization assembliesis that the array of filtrates taken from filtration assembly 1600 canbe placed into two or more crystallization assemblies 2100, allowing forcrystallization under different conditions or methodologies. Forexample, this methodology allows for a temperature study to be performedin which more than one array of solutions is crystallized, each at adifferent temperature. As mentioned earlier, using interchangeable partsin the different assemblies allows for efficiency in inventory and partsuniformity. The different assemblies also permit flexibility in processflow for each part of the recrystallization procedure.

Crystallization assembly 2100 has lower reactor base 2160. Reactor base2160 has recessed region 2161 that is constructed to receive substrate2164, which is resting on top of pad 2162. Pad 2162 should be selectedto help avoid breaking the substrate. This pad is preferably foam,silicone rubber, or another resilient material.

Substrate 2164 provides a suitable surface for crystal formation.Moreover, substrate 2164 can be transferred to the desired analyticalequipment to enable characterization of the array of crystals. Thesubstrate may be constructed from an optically transparent material,such as glass, to allow for direct optical studies of the crystals onthe substrate. It can also be made of other materials such as plastic.Substrate 2164 may have a substantially flat surface on which thecrystals are contained. Substrate 2164 may have distinct regions on itssurface to support crystals formation. For example, those distinctregions may be round, square, etc. The regions may also be recessed toreadily retain crystals. Thus, dimples or wells can be used. (A dimpleherein is defined as a depression with a rounded concave surface, and awell is a recess with one or more defined sidewalls.)

Above substrate 2164 is gasket 2166 having an array of holes (not shown)that correspond to the array of regions on substrate 2164. Whenassembled, these lower components (pad 2162, substrate 2164, and gasket2166) will fit into recessed region 2161 so that gasket 2166 is securedsubstantially flush against the upper outside edges of reactor base2160. Crystallization reactor 2168 is placed over these components andsecured to reactor base 2160 using bolts 2170 or other securing means(clips, clamps, nuts, etc.). The securing means should providesufficient force to compress gasket 2166, which in turn seals theregions on substrate 2164 from potential crosstalk.

Crystallization assembly 2100 also includes sheet 2172 (typically madeof Teflon®) and septum 2174 (which may be identical to barrier sheet2118 and septum 2120, respectively) and reactor cover 2176 coveringcrystallization reactor 2168. Reactor cover 2176 is similar to reactorcover 2122 for reaction assembly 1500, having holes through which needle2020 may pass. The parts over crystallization reactor 2168 are securedto crystallization reactor 2168 by bolts 2178 (although other clamping,clipping or securing means may be used). Thus there are two sets ofsecured components in crystallization assembly 2100. The first setincludes crystallization reactor 2168, gasket 2166, substrate 2164, pad2162, and reactor base 2160. The second set includes sheet 2172, septum2174, and reactor cover 2176, which together are coupled tocrystallization reactor 2168. These separate sets permit the bottom partof crystallization assembly 2100 to be assembled without the upperportion, resulting in more flexibility in being able to use part or allof crystallization assembly 2100 for pre-formulation testing.

Reactor cover 2176 and septum 2174 may not be used if evaporationstudies are to be tested with the material samples contained withincyrstallization assembly.

FIG. 21A illustrates an alternative embodiment of a crystallizationassembly 2102 that is designed to permit formation of crystals inaccordance with the principles of the present invention. Crystallizationassembly 2102 includes similar parts as crystallization assembly 2100such as reactor base 2160, substrate 2164, and reactor 2168. Notincluded with crystallization assembly 2102 is pad 2162 and gasket 2166.Instead substrate 2164 is positioned directly on top of reactor base2160 and o-rings 2104 are added to crystallization assembly 2104.O-rings 2104 may be comprised of a sheet of o-rings (as shown in FIG.21B) or it they can be a two-dimensional arrays (e.g., 4×4 array ofindividual o-rings). When crystallization assembly 2102 is assembled,o-rings 2104 are pressed flush against substrate 2164 and reactor 2168to provide effective isolation of material samples deposited into thecrystallization assembly 2102.

FIG. 21C is a partial cross-sectional view of crystallization assembly2102 that illustrates how the through-holes of reactor 2168 andrespective portions of substrate 2164 are isolated from each other byo-rings 2104. Also shown in FIG. 21C are the wells 2106 in which o-rings2104 reside. Thus when substrate 2164 is pressed against reactor 2168,o-rings 2106 are pressed against substrate 2164 and wells 2106, therebyproviding a substantially airtight seal that isolates each region of thesubstrate.

When aspirating from or dispensing material into a sealed vial through aseptum using a needle or similar device, the accuracy in controlling thevolume of material transferred can be affected by the changing volumeand pressure within the vial. One solution to this problem has been touse a coaxial needle, which consists of two tubes attached togetheralong their axises. One tube is used to transfer material to or from thevial, and the second tube is used as a vent to equilibrate pressurebetween the vial interior and the surroundings.

The coaxial needle device presents several shortcomings. Because thetwo-tube assembly is much larger than a single needle, it will createlarger holes when piercing a septum that may not seal properly when theneedle is removed. Also, because the tips of the tubes remain at a fixeddistance with respect to one another, the range of depths in the vial towhich material can be introduced or withdrawn is limited. Dispensing alarge amount of material into a deeper vial, for example, may lead tomaterial flowing out of the vent tube if the tip of the vent tubebecomes submerged. These shortcomings of the prior art are addressed bythe venting needle device described in conjunction with FIGS. 22A and22B.

FIG. 22A shows a venting needle assembly 2200 that can be used toaspirate from, or dispense materials into, sealed vials or receptacles.Venting needle assembly 2200 comprises venting needle 2202, liquidhandling needle 2204, body 2206, guide rods 2208, springs 2210, andguide block 2212. Body 2206 is attached to a liquid handling system (notshown) that controls aspiration and dispensation of liquid throughliquid handling needle 2204. Liquid handling needle 2204 extends a fixeddistance from body 2206 and passes through movable guide block 2212.Venting needle 2202 also passes through movable guide block 2212,essentially parallel to liquid handling needle 2204, and extends a fixeddistance from the lower face of guide block 2212. Guide block 2212 isattached to guide rods 2208, and configured so that it can slide alongthe central axis of liquid handling needle 2204. Guide rods 2208 areconfigured to slide into body 2206 when force is applied to the lowerface of guide block 2212. Springs 2210 are located on guide rods 2208,between body 2206 and guide block 2212. These springs return guide block2212 to a fully-extended position, determined by the length of guiderods 2208, when there is no force on the lower face of guide block 2212.

As described above, reactor assembly 1500, filtration assembly 1600, andcrystallization assembly 2100 each include a sealing layer (e.g.,septum) for isolating each vial or receptacle from each other. Forpurposes of brevity and clarity, venting needle assembly 2200 isdescribed in conjunction with reactor assembly 1500. Venting needleassembly 2200 is not limited to being used only with reactor assembly1500, but it can be used with any suitable assembly comprising sealedvials or receptacles. If desired, venting needle assembly 2200 can beused in place of pipette system 2060.

After reactor assembly 1500 is sealed, each vial contains a fixed volumeof material. Venting assembly 2200 prevents the pressure from changingwhen aspirating liquid from or dispensing liquid into a sealed vial,which permits increased accuracy in controlling the volume of materialtransferred. Venting assembly 2200 accomplishes this by using a dualneedle assembly, wherein one needle is used to aspirate/dispense and theother is used to vent the vial.

In a typical material transfer process, venting assembly 2200 is loweredover reactor assembly 1500, either manually or by mechanical means,until the lower surface of guide block 2212 contacts the upper surfaceof reactor cover 1522. Both venting needle 2202 and liquid handlingneedle 2204 extend far enough from guide block 2212 to extend throughholes 1523 above the vial of interest, and pierce septum sheet 1520 andbarrier sheet 1518 (if present). When venting needle 2202 pierces septumsheet 1520 (and barrier sheet 1518, if present), it provides an openpassageway between the upper portion of the vial and the surroundingatmosphere, which maintains equilibrium between the pressure in the vialand the ambient pressure (i.e., pressure outside the sealed vial). Thus,when liquid is aspirated from or dispensed into the vial by liquidhandling needle 2204, the pressure does not change because ventingneedle 2202 provides the substantially constant pressure with the vial.

Body 2206 may be pushed towards the top of reactor assembly 1500 untilliquid handling needle 2204, which is attached to body 2206, reaches thedesired depth within the vial for dispensing or aspirating a liquid.Body 2206 may contain a mechanism (not shown) the heat liquid handlingneedle 2204 to a predetermined temperature. As liquid handling needle2204 extends into the vial, guide rods 2208 slide further into body 2206and as springs 2210 compress, as shown in FIG. 22B. During thisprocedure, venting needle 2202 extends a fixed depth into the sealedvial, preferably with the tip of venting needle 2202 located near thetop of the vial, above any liquid which may be present. This device thuspermits transfer of a liquid to and from vials having different depthswhile maintaining a venting passage that equilibrates pressure betweenthe interior of the vial and the ambient atmosphere. When the transferof liquid is complete, body 2206 may be pulled away from reactorassembly 1500, causing liquid handling needle 2204 to withdraw from thevial and springs 2210 to expand until guide block 2212 reaches itsfully-extended position, as shown in FIG. 22A. Pulling body 2206 furtheraway from reactor assembly 1500 removes both venting needle 2202 andliquid handling needle 2204 from reactor assembly 1500 through holes1523. The device may then be inserted into another hole over a differentvial and the above procedure repeated.

Temperature is an important parameter in the various formulations,filtration, and crystallization operations that enable salt selectionand polymorph production. For example, in some methodologies it isdesirable to heat reaction assembly 1500 to dissolve as much drugcandidate as possible in the solvent(s). In processes where temperatureis important (e.g., during crystallization), any of assemblies 1300,1500, 1600 or 2100 (shown in FIGS. 13, 15, 16, and 21, respectively) maybe placed in an oven, cooler, or other temperature-controlled chamber(such as the Torrey Pines incubator cited herein).

It may be desirable in some processes to have precise control oftemperature. One method for controlling the temperature of theassemblies during certain stages of the workflow is shown in FIG. 23,which depicts three-axis robot 2300, having arms 2305 and 2310 andplurality of pumps 2334, situated by work surface 2330. Pumps 2334 arein fluid communication with a solvent or other liquid and in fluidcommunication with needles 2320 and 2322. Heated block assembly 2350 ispositioned on work surface 2330. Heated block assembly 2350 isconfigured to contain reaction assemblies 1500, filtration assemblies1600 and those assemblies to assist mixing material samples. It may bedesirable to heat filtration assembly 1600 to prevent crystallizationduring the filtering process. Not shown in FIG. 23 is a magneticstirring device that can tumble magnetic stirring fleas to agitatematerials contained within the assemblies. Also located on work surface2330 are several temperature-controlled housings 2500.Temperature-controlled housing 2500 is described in more detail inconjunction with the description accompanying FIG. 25. Housing 2500 cancontain an assembly such as the reactor assembly 1300, reaction assembly1500, filtration assembly 1600, or crystallization assembly 2100. FIG.23 also shows deck assemblies 2370 that hold various assemblies for usein a workflow.

Detailed structure of the temperature-controlled housing is not criticalto this invention, and may simply comprise insulated walls with aresistive heater regulated by a thermocouple for temperature control.These types of heaters are sold commercially as the FIREROD™ heatersavailable from Watlow of St. Louis, Mo.

FIG. 24 illustrates an exploded view of temperature-controlled housing2350 shown in FIG. 23. Temperature-controlled housing 2350 includes tub2410 and tub support fixtures 2430 and 2440. Tub 2410 is constructedfrom a material (e.g., aluminum) having desired thermal conductiveproperties. As shown, tub 2410 has pockets 2412 and 2414 for containingan assembly (e.g., reactor assembly 1500 or filtration assembly 1600).Tub 2410 has fluid channels (not shown) in which a fluid or gas isprovided to heat or cool the tub. Fluid inlet/outlet ports 2354 provideconnections to allow a desired heating or cooling fluid or gas to flowthrough the channels, thereby heating or cooling an assembly placed inhousing 2350. The temperature-controlled housing shown in FIG. 24 may besurrounded by an insulating layer to improve thermal control andstability.

Another embodiment that facilitates temperature control of a reactor orassembly is shown in FIG. 25. FIG. 25 illustrates thermal controlchamber 2500 having top 2570 and bottom 2580 that mate to form anenclosure. Top 2570 has cavity 2572 therein and bottom 2580 has cavity2582 therein, which together define the enclosure. Cavity 2572 ispreferably sized to accommodate at least one of the assemblies 1300,1500, 1600, or 2100 (shown in FIGS. 13, 15, 16, and 21, respectively).

In a preferred embodiment, top 2570 has holes 2574 to allow forcommunication through top 2570, (e.g., by a needle, cannula or pipette),for aspirating or dispensing liquids into the regions, vials or wells ofthe assembly in the cavity when top 2570 and bottom 2580 are closedtogether. A fluid that heats or cools the thermal control chamber mayflow through top 2570 and/or bottom 2580. The fluid flow preferablymaintains a specific temperature substantially throughout the assemblycontained therein. A temperature variation of about 2° C. across theenclosed assembly is generally acceptable, but a temperature variationof less than about 1° C. is desirable. Top 2570 and bottom 2580 are madeof a material (e.g., aluminum) with good heat transfer properties.Suitable fluids with well-characterized thermophysical properties (e.g.,water, glycol, etc.) flow through top 2570 and/or bottom 2580 forheating or cooling.

Thermal control chamber 2500 can apply a thermal profile to the assemblycontained therein. The thermal profile may, for example, subject theassembly to a range of temperatures that vary with time or to atemperature that is substantially the same for the chosen time periodthroughout the thermal control chamber. For example, the temperature ofthe thermal control chamber may ramp up to a predetermined temperature aspecified rate, stay at the predetermined temperature for fixed periodof time, and then ramp down to ambient temperature after a specifiedperiod of time.

FIG. 25 shows thermal control chamber 2500 wherein top 2570 and bottom2580 are connected by hinge 2586 that allows the top and bottom to pivotwith respect to each other. This configuration allows for easieralignment and fastening of the top and bottom of the thermal controlchamber that contains an assembly such as crystallization assembly 2100(shown in FIG. 21). When top 2570 is pivoted about hinge 2586, it alignswith bottom 2580 so that cavities 2572 and 2582 meet to enclose theassembly within the chamber.

A typical fluid flow path can occur as follows. Thermal fluid enterseither top 2570 or bottom 2580 first and then flows through the inlet,transverse, and outlet conduits. Then fluid passes through externalconduit 2588 and into the other half of the chamber where the flowpattern is repeated. Thus, if the thermal fluid enters top 2570 first,it will flow through the top, exit the top and be channeled into bottom2580, flow through the bottom, and then exit the bottom. The flowdirection may also be reversed if desired, so the thermal fluid entersthe bottom before the top. Rotating fluid transfer hinge 2587 may beprovided on either side of hinge 2586 which allows top 2570 and bottom2580 to pivot with respect to each other without adjusting externalconduits 2588. This function could also be accomplished with flexibleexternal conduits and a loop, however, the arrangement shown in FIG. 25is a preferred embodiment as it avoids the need for flexible tubing.

Although not shown in FIG. 25, chamber 2500 has a window that provides ameans for a device to monitor the arrays of material samples containedwithin chamber 2590. A glass sheet is coupled to this window with agasket. Preferably a purge of dry gas (e.g., nitrogen) is provided toprevent condensation from developing when chamber 2590 cools. Thisensures that the monitoring equipment has an unobstructed access to thematerial samples. For example, in-situ measuring requires anunobstructed access to monitor the materials samples that are subjectedto crystallization conditions.

The structure that facilitates fluid flow is shown in more detail inFIG. 26, which depicts a cutaway view of the channels that define thefluid path in top 2570 and/or bottom 2580 of thermal control chamber2500. The heating/cooling fluid enters via fluid inlet 2590, which leadsto inlet conduit 2592. A plurality of transverse conduits 2594 lead frominlet conduit 2592 to outlet conduit 2598 and to fluid outlet 2599.Inlet conduit 2592 and outlet conduit 2598 are preferably larger indiameter than the transverse conduits 2594 so that the fluid enteringinlet 2590 will flow uniformly through all of transverse conduits 2594.This flow pattern for the heating/cooling fluid assists in uniformheating or cooling of control chamber 2500

Transverse conduits 2594 pass on either side of holes 2574, if present.Larger conduits 2592 and 2598 may be formed by drilling holes through ablock and then plugging one end of each with a large plug 2595 for easeof manufacture. Smaller transverse conduits 2594 can be formed in asimilar manner, with both ends of the drilled holes capped off by smallplugs 2596.

In operation, a thermal fluid such as water, glycol, or glycerol istransported from a uniform temperature reservoir (not shown) to top 2570or bottom 2580 of thermal control chamber 2500 through a fluid lineusing a constant or variable-speed pump. The thermal fluid enters inletconduit 2592 through inlet port 2590. The thermal fluid flows throughtransverse conduits 2594 and into outlet conduit 2598. The thermal fluidthen exits through fluid outlet 2599, where it then returns to thereservoir or is routed through another top or bottom.

A heat pump or other device may be used to regulate the temperature ofthe thermal fluid in the reservoir by adding or removing heat through aheat transfer coil located in the reservoir. In one embodiment, aprocessor receives signals from temperature sensors located in thereservoir, and adjusts the amount of heat added to or removed from thethermal fluid through the coil. One or more temperature sensors (notshown) may also be mounted in the top or bottom of the thermal controlchamber and connected to the processor. Temperature control can then beachieved by varying both the temperature of the thermal fluid in thereservoir and the flow rate through the conduits. To adjust the flowrate of thermal fluid through the fluid flow system, the processorcommunicates with a valve located in a reservoir outlet line. Variousparts of the system can also be insulated to improve temperature controland stability.

FIG. 27 shows a cross-sectional view of thermal control chamber 2500with crystallization assembly 2100 enclosed therein. This view showsinsulation 2591 can be packaged with thermal control chamber.Cross-sectional view 2700 illustrates additional details of thecomponents just described above in conjunction with FIGS. 21, 22, 25,and 26.

Determining the melting point of salts and drug candidates is animportant characterization technique for the screening process. Inparticular, differences in melting point among crystals or amorphousprecipitates of a given compound comprise one of the best indicators fordetermining the existence of polymorphs.

In one embodiment of the present invention, a melting point station isemployed to determine the melting point of the solid material containedin each region of an array following salt formation, precipitation,crystallization, or other similar procedure in the process. Meltingpoints of various materials may be determined by controllably raisingthe temperature of the substrate containing the array while monitoringthe birefringence image (if present) of one or more regions of thearray. The temperature at which the birefringence image from a region ofthe array disappears indicates a transition from the solid to the liquidstate. This transition state corresponds to the melting point of thesolid matter (generally crystals or amorphous solid) in that region.

Generally, this measurement is based on melting point measurements knownin other fields of endeavor. See, e.g., Magill, J. H. “A new method forfollowing rapid rates of crystallization. II. Isotactic Polypropylene,”Polymer 1962, 3, 35-42; Ding, Z. and Spruiell, J. E. “Anexperimental-method for studying nonisothermal crystallization ofpolymers at high cooling rates,” J. of App. Polymer Science PartB—Polymer Physics, 1996, 34, 2783-2804; Garetz, B. A., et al.,“Birefringence and diffraction of light in ordered block-copolymermaterials,” Macromolecules, 1993, 26, 3151-3155; Carlson, E. D. et al.“Mechanical and Thermal Properties of Elastomeric Polypropylene,”Macromolecules, (1998), 31(16), 5343-5351; and Carlson, Eric David, “Arheo-optical investigation of the relaxation and crystallizationbehavior of stereoblock polypropylene synthesized from 2-arylindenemetallocene catalysts,” Stanford Ph.D. Dissertation (1998); each ofwhich is incorporated by reference.

A melting point apparatus designed to determine melting points of anarray of solids on a substrate is shown in FIG. 28. Melting pointapparatus 2800 determines the melting points of an array of samples bymonitoring the optical properties of the samples for phase transitions.Melting point apparatus 2800 can also measure recrystallization kineticsduring thermal cycling. Melting point apparatus 2800 can use a lightscattering or birefringence measuring technique to determine when asample transitions from a crystalline structure to an amorphous liquid.When performing birefringence measuring, a transmissive or a reflectivetechnique can be used.

Apparatus 2800 detects the melting point of material samples as follows.First, a sample array is prepared via crystallization. Those of skill inthe art will appreciate that a sample array can be prepared by othermethods other than crystallization. For example, an evaporation or aprecipitation method can be used. It will also be appreciated that solidmaterials (e.g., may be prepared by hand or automatically by a robot.The sample array may be contained on a glass slide (e.g., universalsubstrate) or other transparent material that is placed into a carrierblock. If desired, the carrier block can be covered to contain anymoisture that forms from products of material decomposition such ascondensation, solvent vapors, etc. Finally, the carrier block is placedinto a temperature chamber such as thermal control chamber 2500 or isplaced within two insulated plates (FIGS. 28A and 28B), which form aquasi-isothermal block.

After the material samples are placed in the temperature chamber, anindividual light source and a detector are aligned to each materialsample in the array. Operating in transmission mode, each light sourcetransmits a light signal through a material sample. The light source canany suitable device such as, for example, light emitting diodes, lasers,or incandescent lights. Collimated, polarized, intense, monochromaticlight is preferable for operation, but weak light (e.g., incandescentlight) can also be used. The light signal can comprise a wavelengthranging between about 300-2000 nanometers (nm). If desired, a singlelight source can be distributed to each material sample by, for example,a fiber-optic bundle, a telecentric lens, a telescope, collimatingoptics, or other similar apparatus. Alternatively, a single light sourceor an array of light sources may be moved across each material sample toilluminate a single material or a row of such materials at a time.Persons skilled in the art will appreciate that any suitable arrangementof light sources can be used to provided light-scattering orbirefringence testing.

If a reflective technique is used, the light path does not pass throughthe material samples. Rather, the reflection of light off a crystallinestructure is detected. This technique is described in detail inconjunction with in-situ measuring assembly 3000 of FIG. 30. Thereflective technique operates generally as follows. Light emitted from alight source passes through a first polarizer, then it reaches a beamsplitter that redirects the light towards a material sample. If desired,the light can be “pre-polarized” (e.g., a laser) so that a firstpolarizer is not needed. Light reflected from the material sample passesback to the beam splitter, but passed through the beam splitter. If thelight signal has been altered (e.g., polarized) by the material sample,then it may pass through a second polarizer that is rotated 90° from thefirst polarizer and detected by a detector.

Light provided by the light source first passes through a polarizingfilter, then through the material sample, through a second polarizingfilter that is oriented at 90° with respect to the first polarizingfilter. If the material sample does not contain a crystalline structure,then the light signal will be extinguished by the second polarizingfilter, thereby preventing a detector from detecting the light signal.If the material sample contains a crystalline structure, however, thecrystal will rotate the polarization of the light, thereby allowing someof the light to pass through the second polarization filter. Any lightthat passes through the second polarization filter is detected by thelight detector.

Various detection methods can be used to detect light emitted by thelight source(s). Individual detectors such as photodetectors (e.g.,silicon photodiode, photovoltaic cell, etc.) may be positioned withrespect to each material sample. Each photodetector is connected to acomputer (e.g., computer 110) that monitors the output of each detector.Based on the output signals of the photodetectors, the computer candetermine at what temperature a crystalline structure melts. Likewise,if a detector such as a camera, a charge-coupled device (CCD) a digitalcamera, or a video camera is used to detect phase transitions, acomputer may register the transitions and calculate the melting pointtemperature.

Initially, before the chamber begins to heat the material samples, eachdetector can ascertain whether a crystal structure is present. Becauseeach detector is connected to a computer or other data logging device,each detector can constantly monitor each material sample. This isuseful because when the material samples are heated, they eventuallymelt, and the detectors detect this phase transition. When the crystalmelts, it becomes amorphous, and light is no longer able to pass throughthe second polarizing filter. When a detector no longer detects apreviously existing light signal, it is at this temperature that thecrystal has melted.

The temperature of the material samples are ramped up at user definedrate to a user defined maximum temperature. The rate of temperatureincrease is precisely controlled and uniform throughout the materialsamples. This enables apparatus 2800 to accurately measure melting pointtemperatures. Alternatively, the temperature can be step up incontrolled increments. Furthermore, the temperature of each materialsample can be controlled individually.

The above discussion was primarily directed to birefringence testing,but the present invention is not limited to such. Differentbirefringence testing can be performed. For example, the polarizationangle of one or both polarization plates can be modulated. The frequencyof the light can be modulated. In another approach, the above mentionedlight scattering approach can be implemented by removing the polarizerplates. Light scattering, for example, is another testing method thatcan be implemented to detect crystalline structures contained in thesample array.

Apparatus 2800 includes two main assemblies: thermal platform 2810 thatcan support and controllably heat and cool an array (i.e., a substratecontaining samples), and opto-mechanical platform 2830 that permitssimultaneous optical imaging of some or all of the regions on the array.

Thermal platform 2810 includes thermal platform top 2812 and thermalplatform base 2814, which when assembled together create cavity 2816that supports a substrate or suitable testing platform (e.g., a carrierplate). Thermal platform 2810 is made of any suitable material that hashigh thermal conductivity such as aluminum, aluminum alloys, or copper.Other materials such as indium, lead, tin, or silver may be used to formpart or all of thermal platform 2810 to enhance conductivity and improvetemperature uniformity. The substrate may be constructed fromborosilicate glass, or is a Zinnser™ plate, or is made of some otheroptically transparent or translucent material. The substrate ispreferably positioned to provide optimal thermal conduction with thermalplatform 2810. For example, substrate 2164 (FIG. 21) can be placed inthermal platform 2810.

Thermal platform 2810 is shown in detail in FIGS. 28A and 28B. FIG. 28Ashows an isometric view of melting point apparatus 2800 (computer notshown) that shows thermal platform 2810 and light source array 2832. Asdescribed above, thermal platform 2810 includes thermal platform top2812 and thermal platform bottom 2814, which are also shown in FIG. 28A.Thermal platform 2810 is shown to be in an OPEN position. That is,pneumatic press 2860 is in a position such that thermal platform top2812 and thermal platform bottom 2814 are not pressed against eachother. Pneumatic press 2860 is used to assist opening and closingthermal platform 2810.

FIG. 28B shows a cross-sectional view of the FIG. 28A taken along line28-28. Thermal platform 2810 is also shown to be in the OPEN position.One skilled in the art will appreciate that when thermal platform 2810is closed, thermal platform top 2812 and thermal platform bottom 2814are pressed flush against one another to fully enclose substrate 2864contained therein. Preferably, a glass cover sheet is placed oversubstrate 2864 to contain any products of composition (e.g.,condensation, out-gassing).

Both thermal platform top 2812 and thermal platform bottom 2814 havethrough holes 2823 in which resistive heating elements can be contained.Theses resistive heating elements, which are described in more detailbelow, provide a mechanism for heating substrate 2864 at a controlledrate while maintaining a uniform temperature distribution.

Referring back to FIG. 28, thermal platform base 2814 is designed to beheated by elements attached to or embedded within thermal platform base2814. In a preferred embodiment, thermal platform base 2814 containschannels (not shown) that are similar to conduits 2592, 2594, and 2598in FIG. 26. A thermal fluid from a reservoir (not shown) can flowthrough these channels to heat the substrate in a controlled manner, asdescribed above. A thermal fluid at a lower temperature can also flowthrough these channels or an adjacent set of such channels to coolthermal platform base 2814.

Alternatively, one or more resistive heating elements 2822 may beattached to or embedded in thermal platform base 2814 to provide amechanism for heating thermal platform base 2814. Resistive heatingelements 2822 may include, for example, wire-wound resistive heaters,thermoelectric devices (e.g., peltier junctions) and thin- or thick-filmresistor heaters. Resistive heating elements 2822 may be embedded orattached to thermal platform base 2814 to permit improved control ofthermal uniformity or specific temperature profiles in thermal platform2810. Different resistive heating elements 2822 may be capable ofdistributing different quantities of heat (i.e., the power consumptionmay vary). Various resistive heating elements 2822 that have differentpower consumption may be placed strategically on thermal platform top2812 and thermal platform base 2814 to promote uniform temperaturedistribution. A combination of channels for heating/cooling by flow ofthermal fluid and resistive heating elements 2822 may also be used.Those skilled in the art will recognize that other methods of heating orcooling thermal platform base 2814 in a controlled manner may also beused.

Another method for heating or cooling the material samples is to subjectthe samples to a gaseous bath such as that provided by a convectionoven. Yet another method is to heat each material sample individually,or in a serial fashion with individual heaters.

Thermal platform 2810 may also have one or more embedded or attachedthermal sensors 2820 that can be used in connection with appropriateexternal equipment (e.g., datalogger, computer, etc.) to monitor thetemperature of the platform. These sensors may be in the top or bottomof platform 2810, and may include one or more of the following:thermocouples, resistance temperature detectors (RTD's), orsemiconductor-based thermistors. One or more of these sensors may belocated either in the top, in the bottom, or in both the top and bottomof thermal platform 2810. Signals obtained from these sensors arepreferably connected to a computer (e.g., computer 110 of FIG. 1) topermit automated monitoring and control of the temperature. The sensorsignals can be processed using appropriate control software to regulatecurrent in resistive heater elements, if present, or to regulate thetemperature and flow rates of thermal fluids in channels, if present, orto provide more accurate control of both heating and cooling rates inthermal platform 2810.

Various sensor arrangements can be used to accurately determine themelting point of crystalline structures. The temperature of eachmaterial sample may be individually measured, interpolated from an arrayof neighboring sensor, or extrapolated from a single sensor. Sensors maybe located on thermal platform 2810 or some other heat transfer medium(e.g., convection oven). Sensors may be located directly on the samplesubstrate such as directly below a well containing a material sample.

Thermal platform 2810 implements a heating system that is capable ofraising the temperature of thermal 2810 platform and an enclosedsubstrate from ambient temperatures to a user defined temperature. at auser defined rate. For example, the user can select the maximumtemperature to be about 200° C., 280° C., 300° C., or any other suitabletemperature. The rate at which the temperature can be increased ordecreased can also be set by the user. For example, a user may set therate of temperature deviation at about 0.5° C. per minute, 1.0° C. perminute, 2.0° C. per minute, or some other suitable temperaturevariation. Persons skilled in the art will appreciate that thermalplatform 2810 may heat or cool an enclosed substrate to any suitabletemperature. In addition, thermal platform 2810 may heat or cool anenclosed substrate at any rate. Moreover, thermal platform 2810 maycycle the temperature to perform re-crystallization studies.

Thermal platform top 2812 is constructed with an array of upper opticalholes 2813 that corresponds to an array of lower optical holes 2815 ofthermal platform base 2814. Holes 2813 and 2815 are similar to holes2574 in thermal control chamber 2560 shown in FIG. 25, are located aboveand below some or all of the regions on a substrate when the substrateis placed in thermal platform 2810. Holes 2813 and 2815 are associatedwith a region of a substrate are constructed to provide an opticalpathway that passes through thermal platform 2810. This optical path ispreferably in a direction substantially perpendicular to the enclosedsubstrate, and intersects at least a portion of the region of thesubstrate associated with the upper and lower holes.

Opto-mechanical platform 2830 in FIG. 28 includes light source array2832 and sensor array 2834 arranged on opposite sides of thermalplatform 2810. Opto-mechanical platform 2830 may be similar to thattaught in U.S. Pat. No. 6,157,449, which is incorporated herein byreference in its entirety. Light source array 2832 may includes one ormore laser diodes, light emitting diodes (LEDs), or other controlledlight sources 2833. Alternatively this array may comprise one or morelight sources that are segregated by mirrors or prisms to form an arrayof beams. Sensor array 2834 includes a group of sensors 2835 such asphoto diodes, charge-coupled devices (CCDs) or other optical sensors,which are preferably positioned on the side of thermal platform 2810that is opposite light source array 2832. First polarizing sheet 2840,typically a commercially-available polarizing filter or polarizingmirror, is mounted between the light source array and the thermalplatform. Second polarizing sheet 2842 is mounted between thermalplatform 2810 and sensor array 2834 at a cross-polarizing orientation ofapproximately 90° with respect to first polarizing sheet 2840.

If desired, the orientation of second polarizing sheet 2842 may not bepositioned exactly at 90° with respect to first polarizing sheet 2840.Rather, second polarizing sheet 2842 may be positioned a few degrees(e.g., 1-5 degrees) above or below a perfect 90° cross-polarization.This slight deviation from perfect cross-polarization preventsextinction of unaltered light signals provided by a light source. Butthis deviation still allows for detecting changes in crystals using abirefringence monitoring technique and/or a light scattering monitoringtechnique.

Light source array 2832, light sensor array 2834, polarizing sheets 2840and 2842, and thermal platform 2810 are configured so that optical path2850 exists between one or more of the light sources and opposing lightsensors that passes through polarizing sheet 2840, thermal platform 2810including a region on the substrate mounted therein, and secondpolarizing sheet 2842. The orientation of second polarizing sheet 2842filters or blocks nearly all of the light that originates from lightsource array 2832 and traverses optical path 2850 without passingthrough any materials. Thus, if light passing through a material and hasits polarization changed, it may pass through second polarizing sheet2842 and be received by light sensor array 2834. If this occurs, thenthe birefringence image of the material (e.g., crystal) may be obtained.

The melting point apparatus described above permits detection of phasetransitions, including the solid-to-liquid transformation, that mayoccur on individual wells or regions of a substrate. A substratecontaining deposits of candidate salts, neutrals, solid precipitates, orfiltrates to be characterized may be located on one or more regions ofthe substrate. This substrate may be placed into thermal platform 2812,which in turn is placed into opto-mechanical platform 2830 to performoptical scanning. Thermal platform 2810 is configured so upper and loweroptical holes 2813 and 2815 align with optical paths 2850. Thisoperation may be performed manually or automatically using robot arms toprovide for a more fully automated procedure which reduces thepossibility of errors in handling the samples.

The temperature of the thermal platform containing the substrate and oneor more material samples is then increased at a defined rate andmonitored. In a preferred embodiment, one or more temperature sensors incontact with the substrate are used to provide real-time temperaturedata to an external data collection and processing device such ascomputer (e.g., computer 110 of FIG. 1). The computer may also beinterfaced to the heating/cooling system described above to providefeedback for better temperature control. In a typical process, thetemperature of the thermal platform is raised from ambient temperaturesto about 200° C., or from ambient temperatures to about 300° C., at arate of about 0.5° C. per minute, 1.0° C. per minute, 2.0° C. perminute, or any other suitable rate.

The optical signal received by each light sensor 2835 is monitored by acomputer in real time. Transformation of a material candidate or othermaterial on a region of the substrate from a solid to a liquid statewill generally cause an abrupt decrease in the amount of light reachingthe sensor that lies on the optical path passing through that region orwell. This effect arises primarily because amorphous liquids have a muchweaker “birefringence polarizing effect” on light than solid crystallinematerials. The temperature at which the optical signal associated with agiven region or well of the substrate changes can then be detected bythe computer and recorded, yielding a measure of the melting point ofthat particular sample. The entire array of materials on the substrateor any subset thereof may be characterized for melting points during asingle thermal ramp-up cycle, as the individual sensor corresponding toeach region will provide a distinct signal as the temperature of theentire substrate reaches the melting point associated with the materiallocated at said region.

Slower rates of temperature increase tend to yield more accurate meltingpoint values by reducing transient errors associated with finite heattransfer delays between the thermal platform and the substrate. Theactual ramp-up rate chosen for a given measurement set will generallyreflect a compromise between accuracy and overall time required tocomplete the set.

In the case of polymorphs, it is possible to measure more than simplemelting points with this test, inter-polymorph transitions may also beobserved. Inter-polymorph transitions may include transitions from onepolymorph to another (e.g., solid to solid transitions). In this case, adetectable shift in the intensity of filtered light reaching a sensor(where the change may be an increase or a decrease) would indicate atransition from one solid form to another, particularly if the sameregion of the substrate undergoes a melting transformation at some latertime and higher temperature during the set of measurements.

FIG. 30 illustrates an in-situ monitoring assembly 3000, which can beused to observe crystallization of library members in accordance withthe principles of the present invention. As shown, in-situ monitoringassembly 3000 is positioned below work surface 2330 of FIG. 23, whichprovides support for devices such as thermal control chambers 2500. Inthis configuration, work surface 2330 has optical pathways leading tothe regions (or vessels) where materials are being formed so thatin-situ monitoring device 3000 can perform measurements. As furthershown, assembly 3000 is capable of moving horizontally with respect towork surface 2330 for positioning under various thermal control chambers2500. Assembly 3000 is positioned beneath one of thermal controlchambers 2500 to perform in-situ measurement. If desired, assembly 3000can be positioned above work surface 2330 or part of work surface 2330.In-situ measurements may provide an advance indication of librarymembers that have formed crystal structures. This information can beprovided, for example, as input for determining which library membersshould be selected for testing when the crystallization step iscomplete.

Also shown in FIG. 30 is an enlarged cross-sectional view taken fromcircle 3. The cross-sectional view shows how in-situ monitoring can beaccomplished using light source 3010, diffuser 3012, polarizer 3014,light beam splitter 3016, second polarizer 3018, lens 3020, and detector3022. When assembly 3000 is it preferably positioned such that thecrystallization assembly (e.g., crystallization assembly 2100) containedwithin thermal control chamber 2500 is within full view of lens 3020.This enables assembly 3000 to scan two or more library memberssimultaneously. If lens 3020 is a telescentric lens, then light receivedby the lens is able to transmit the light in parallel to detector 3022,thereby reducing image distortion caused by conventional lenses. Whendetector 3022 captures the light signal, it is able to provide arelatively clear picture of the material samples. Detector 3022 can be,for example, a camera, a digital camera, a television camera, acharge-coupled device.

Assembly 3000 operates on substantially the same principles as thatpreviously described in conjunction with the birefringence testingmechanism of FIG. 28. Instead of a transmission scanning technique,however, assembly 3000 uses a reflective optical scanning technique thatdetects reflection of light off a crystalline structure. The reflectivetechnique operates generally as follows. Light emitted from light source3010, as indicated by the dotted line, passes through diffuser 3012 andfirst polarizer 3014. Then the light signal reaches beam splitter 3016,which redirects a portion of the light signal towards the materialsamples, as indicated by the change in the illustrative light path.Light reflected from the material samples passes back through beamsplitter 3016. If any portion of the light signal has been altered byany of the material samples, then it may pass through second polarizer3018 that is rotated 90° from the first polarizer. Light that passesthrough second polarizer 3018 is received by lens 3020 and then capturedby detector 3022.

Detector 3022 may capture an image of the entire array of materialsamples or it may capture a portion. If it captures a portion of thearray, assembly 3000 may be moved accordingly so that a complete pictureor detection of the array can be obtained.

Use of assemblies 1500, 1600 and 2100 allow for a complete workflow inthe formulation of crystals of drug candidates. In one embodiment, theworkflow begins with the original drug candidate being dispensed intovials 1516 of reaction assembly 1500, either while vials 1516 are in aseparate rack or in reactor base 1514 (e.g., with bottom plate 1510 andshock-absorbent layer 1512 attached to reactor base 1514). The drugcandidate can be in a solid state or in solution or suspension, but anysolvent present with the original form of the drug candidate is removed,for example, by evaporation, wicking, or other methods known to those ofskill in the art. The desired recrystallization solvent or solventmixtures selected as discussed above are then dispensed into each vial1516 in the desired amounts (typically with sufficient solvent to formsaturated solutions in the vials). Optionally, different acids, bases,or salts are added (as discussed above in the salt selection process).Also optionally, mixing objects are placed in the vials (for example,using device 1402 such as in FIG. 14).

Barrier sheet 1518 and septum 1520 are placed over vials 1516 in reactorbase 1514 (with bottom plate 1510 already attached) and reactor cover1522 is secured to reactor base 1514 with sufficient strength to formseals over vials 1516. Assembled reaction assembly 1500 is then placedin a heater, shown in FIG. 23 (or FIGS. 24 and 25) and optionally placedon a commercially available shaker (available through VWR and made byIKA, MTS, WORKS or Lab-line). For a desired amount of time (such as 2hours or more, 4 hours or more or 8 hours or more) and at a desiredtemperature (such as at least 40° C., at least 60° C. or at least 80°C.), assembly 1500 is heated and optionally stirred or shaken to allowfor dissolution and/or reaction.

Afterwards, reaction assembly 1500 is placed on a work surface and aneedle or pipette is used to sample the hot liquids in vials 1516. Thisis accomplished by inserting a needle or cannula through holes 1523,septum 1520 and barrier sheet 1518 and aspirating an aliquot of liquid(such as less than 1000 μL or less than 100 μL). The aliquot can betaken by hand or automatically, such as with the equipment shown in FIG.23A as described above.

The aliquot of liquid is maintained in the needle or pipette that ismoved to filtration assembly 1600, which is in an assembled state. Theneedle is placed into the first position as shown in FIGS. 20A and 20B,and extended through hole 1625A, septum 1620 and barrier sheet 1618 suchthat it is in sealing communication with small o-ring 1744. The liquidis dispensed through needle 2020 into opening 2020 and the liquid isfiltered through the filter 1634 before entering vials 1616. Filteringcan occur at a desired temperature by placing filtration assembly 1600into a heater such as described above in FIG. 23.

After filtration, the filtrates (typically at a desired temperature) areaspirated by placing needle 2020 through hole 1625B, as shown in FIG.20C. Needle 2020 extends through hole 1625B, septum 1620, barrier sheet1618, filter subassembly 1630 and into vials 1616 to aspirate an aliquotof the liquid filtrate (e.g., less than 1000 μL or less than 100 μL ofthe filtrate). This filtrate is used for solubility at temperaturetesting, as described herein.

The needle is then moved to crystallization assembly 2100. The needle isextended through holes 2177 in reactor cover 2176 through septum 2174,barrier layer 2172 and into the crystallization receptacle formed byside walls 2169. Liquid filtrate is then dispensed into the receptacle.Liquid filtrate can be dispensed to a number of differentcrystallization assemblies, such as multiple crystallization assemblies,glass microtiter plates and the like so that crystallization occursunder a number of different conditions or methods. After dispensing intoall receptacles, the crystallization assembly is subjected tocrystallization conditions, such as by lowering the temperature of theassembly (e.g., below about 35° C. or below about 25° C. or below about15° C. or below about 5° C.) for a desired amount of time (e.g., twohours or more, four hours or more or eight hours or more).

After the allotted time, reactor cover 2176, septum 2174, and barriersheet 2172 are removed. In an alternative embodiment, thecrystallization assembly can be used without reactor cover 2176, septum2174 and barrier sheet 2172, with the filtrate being deposited directlyinto the crystallization chambers; with this embodiment allowing forother crystallization methodologies, such as evaporation. With reactorcover 2176 removed, the mother liquor or supernatant is sampled forsolubility testing, as described herein. Thereafter, the remainder ofthe mother liquor or supernatant is removed by pipetting and/or wickingor other methods and crystallization assembly 2100 is disassembledproviding an array of crystals on substrate 2164. This array can then bescreened, as described herein. See Example 6 for an illustration ofusing the assemblies described herein in a crystallization workflow.

Although the above-described invention has been described for drugcandidate compounds, this invention may be practiced with any compoundof interest, particularly low molecular weight compounds. Thus, themethods, apparatus and systems described herein may be used for anycompound for which salt selection and/or polymorph characterization isdesired.

The following examples provide illustrative examples on how the presentinvention can be used to perform the processes described above. Theseexamples are for the purpose of illustration only and are not to beconstrued as limiting the scope of the invention in any way.

Example 1 Caffeine—Salt Selection

A solution of caffeine in dichloromethane (25 mg/mL) was dispensed intoninety-six wells of an eight by twelve microtiter plate reactor havingremovable vials (shown in FIG. 15) that may be 750 μl in size such thateach well contained about 10 mg of caffeine. After removal of thesolvent by evaporation, seven different solutions of acids in eitherdichloromethane or tetrahydrofuran (THF) (the salt reactants) weredispensed, with a different acid solution being dispensed into each ofthe twelve wells in different rows of the microtiter plate and with thefirst row having no acid added (only the solvent dichloromethane).

The following acid solutions were added to the twelve wells of each rowstwo through eight: row two, acetic acid in dicholomethane; row three,benzene sulfonic acid in dicholomethane; row four, hydrochloric acid indichloromethane; row five, methyl sulfonic acid in dicholomethane; rowsix, succinic acid in THF; row seven, tartaric acid in THF; and roweight, toluene sulfonic acid in dichloromethane. One equivalent of acidwas added to each well and the total volume was 400 μl. The reactor wassealed and shaken at room temperature for four hours. The cover wasremoved and the solvents evaporated. Twelve different recrystallizationsolvents were added to the wells, with a different recrystallizationsolvent being added to the eight wells of each twelve different columns,as follows: column one, ethylacetate; column two, ethanol; column three,methylethylketone; column four, nitromethane; column five, heptane;column six, aectonitrile; column seven, 2-propanol; column eight,p-dioxane; column 9, 2-methoxyethyl ether; column ten, 1-propanol;column eleven, toluene; and column twelve, water.

The reactor was sealed and heated to 60° C. and heated at thattemperature for four hours. The cover was removed and aliquots (e.g.,200 μl) of each well were removed by pipette and dispensed to a glassmicrotiter plate (from Zinsser Analytics). Another aliquot was removedand added to an array of vials and diluted with acetonitrile for furtherdilution and liquid chromatography analyses, as described above in orderto obtain a solubility measurement of the compound at 60° C. The glassmicrotiter plate was sealed and placed in a Torrey Pines incubator at70° C. The temperature was ramped over 8 hours to 10° C. After sittingat 10° C. for at least ten hours, aliquots of the mother liquor wereremoved and diluted with acetonitrile for LC analysis, as describedabove. The concentration of the caffeine in the mother liquor isconsidered to be the solubility, with the results shown in Table 4,below:

TABLE 4 0.00 4.88 14.54 25.96 0.00 17.36 3.98 18.57 10.88 4.45 4.7426.72 0.00 4.77 12.77 25.67 0.00 16.58 4.03 18.43 10.26 4.55 5.85 26.148.13 4.82 14.54 25.21 0.00 18.82 3.97 19.94 11.12 4.28 5.21 25.38 7.655.25 14.88 18.74 0.00 12.96 3.90 13.70 11.13 4.19 4.96 25.04 4.17 5.199.26 24.28 0.00 16.33 3.58 3.34 2.88 4.22 2.65 25.21 5.29 4.96 14.3725.58 0.00 16.72 4.07 21.53 11.31 4.45 4.54 24.49 2.96 5.58 11.35 18.350.00 5.63 5.05 25.40 12.45 5.19 4.53 25.16 0.17 5.38 7.50 25.90 0.003.52 3.78 3.71 0.94 4.48 0.47 25.66

Note that the solubility measurements at 60° C. and 10° C. can becompared to obtain the mass of the crystals obtained, which was obtainedin this case.

The glass plate was then placed between cross polarizing filters andscanned for wet birefringence. The remaining mother liquor was removedby pipette and residual solvent was removed by wicking with filterpaper. Another image was taken between cross polarizing filter, givingdry birefringence. Birefringence images of the solids in wells wereobtained under a microscope equipped with crossed polarized filters.Raman spectra were obtained on individual crystals in each of the wells,in accord with the procedures described above.

Example 2 Naproxen—Salt Selection

A set of experiments, similar to Example 1, was carried out usingnaproxen to form salts by reaction with bases. The experimental set upwas the same as in Example 1, using a solution of naproxen indichloromethane (25 mg/mL) dispensed into 96 wells such that each wellcontained 10 mg of naproxen. After removal of the solvent byevaporation, seven different solutions of bases in either methanol orwater (the salt reactants) were dispensed, with a different basicsolution being dispensed the twelve wells of different rows of themicrotiter plate and with the first row having no base added (only thesolvent methanol).

The following basic solutions were added to the twelve wells of eachrow: row two, sodium hydroxide in methanol; row three, potassiumhydroxide in methanol; row four, calcium carbonate in water; row five,ammonium hydroxide in methanol; row six, ethylenediamine in methanol;row seven, L-arginine in methanol; and row eight, pyridine in methanol.One equivalent of base was added to each well and the total volume was400 μl.

The reactor was sealed and shaken at room temperature for four hours.The cover was removed and the solvents evaporated. Twelve differentrecrystallization solvents were added to the wells, with a differentrecrystallization solvent being added to the each of the eight wells ofeach different column, as follows: column one, isopropyl acetate; columntwo, ethanol; column three, heptane; column four, acetonitrile; columnfive, 1-octanol; column six, anhydrous p-dioxane; column seven, toluene;column eight, 2-butanone; column nine, water; column ten, nitromethane;column eleven, 1,2-dichloroethane; and column twelve, triethylamine. Thereactor was sealed and heated to 60° C. for four hours. The cover wasremoved and aliquots (e.g., 200 μl) were removed by pipette anddispensed to a glass microtiter plate (from Zinsser Analytics).

Another aliquot was removed and added to an array of vials and dilutedwith acetonitrile for further dilution and liquid chromatographyanalyses, as described above in order to obtain a solubility measurementof the compound at 60° C. The glass microtiter plate was sealed andplaced in a Torrey Pines incubator at 70° C. The temperature was rampedover eight hours to 10° C. After sitting at 10° C. overnight, aliquotsof the mother liquor were removed and diluted with acetonitrile forsolubility at 10° C. by liquid chromatography analysis, as describedabove. The solubility of each of the salts in 1-octanol and water atboth 10° C. and 60° C. were obtained using LC and plotted and used tocalculate the partition coefficient, which is expressed as log P andshown below in Table 5 (pH values were not obtained):

TABLE 5 log P Values Temperature Temperature Cation 10° C. 60° C. None1.51 1.64 Sodium −0.94 −0.77 Potassium 0.28 −0.22 Calcium 0.68 0.78Ammonium −0.57 −0.50 Ethylenediamine −1.05 −0.84 L-Arginine −0.74 −0.85Pyridine 0.82 1.45

The values of this experiment in Table 5 can be compared to the Log Pvalue published in the Physicians Desk Reference for Naproxen of 1.6-1.8at pH of 7.4. This experiment demonstrates that clearly different saltforms of Naproxen were created during this example.

The glass plate was placed between crossed polarizing filters andscanned for wet birefringence. The remaining mother liquor was removedby pipette and residual solvent was removed by wicking with filterpaper. Another image was taken between crossed polarizing filters fordry birefringence.

Example 3 Phenylbutazone—Polymorph Study

A solution of phenylbutazone in dichloromethane (25 mg/mL) was dispensedinto wells of a microtiter plate having removable vials (shown in FIG.3) that were 750 μl in size such that each well contained 10 mg ofphenylbutazone. After removal of the solvent by evaporation, 16recrystallization solvents were dispensed into the wells in the ratiosshown below in Table 6, such that the total volume was 600 μl of solventgenerating a library that included 84 unique solvent compositions. Formixtures of solvents, pure solvents were dispensed prior to mixing andmixed in the wells. The ratios shown in Table 6 are v/v ratios, thus forexample 80/20 means 80 parts of the first listed solvent and 20 parts ofthe second listed solvent (i.e., 480 μl/120 μl). The recrystallizationsolvents were water (W), heptane (H), ethanol (E), dichloroethane (D),acetonitrile (A), methylethylketone (K), toluene (T), dimethylsulfoxide(S), 1-propanol (IP), nitromethane (NM), α,α,α-trifluorotoluene (FT),2-propanol (2P), p-dioxane (I), 1-octanol (1O), propylacetate (PA), andcyclohexane (CH):

TABLE 6 1 2 3 4 5 6 7 8 9 10 11 12 100 80/20 60/40 40/60 20/80 100 10080/20 60/40 40/60 20/80 100 E E/W E/W E/W E/W W H D/T D/T D/T D/T T AA/W A/W A/W A/W W H H/FT H/FT H/FT H/FT FT 1P 1P/W 1P/W 1P/W 1P/W W HH/D H/D H/D H/D D I I/W I/W I/W I/W W H H/PA H/PA H/PA H/PA PA K K/E K/EK/E K/E E D D/A D/A D/A D/A A NM NM/E NM/E NM/E NM/E E D D/S D/S D/S D/SS H H/E H/E H/E H/E E D D/2P D/2P D/2P D/2P 2P 10 10/E 10/E 10/E 10/E ED D/CH D/CH D/CH D/CH CH

The reactor was sealed and heated to 60° C. for four hours. The coverwas removed and aliquots (e.g., 200 μl) were removed by pipette anddispensed into individual regions of a glass microtiter plate (fromZinsser Analytics), creating an array of ninety-six liquids. The Zinsserplate was sealed in the crystallizer and placed in a Torrey Pinesincubator at 70° C. The temperature was decreased to 10° C. over aneight hour time period. After sitting at 10° C. overnight, aliquots ofthe mother liquor were removed and diluted with acetonitrile todetermine the solubility of phenylbutazone in the various solvents at10° C. by LC analysis.

The remainders of the solvents were removed by pipette and any residualsolvent was removed by wicking with filter. The glass plate was thenplaced between cross polarizing filters and scanned for drybirefringence were obtained under a microscope at 5× equipped withcrossed polarized filters. Raman spectra were obtained on individualcrystals in each of the wells in an automated manner, as describedabove. Selected spectra that show the differences between polymorphs areshown in FIG. 9. All of the Raman spectra collected were correlated andgrouped using the software described herein. FIG. 11 shows the spectraby well in array format 2008 as well as the user inputs into thesoftware of minimum grouping correlation coefficient (with suggestedparameters in parentheses) 2002 and whether to use a fixed reference(input of 1) or to use the first well as the reference (input of 0)2004. FIG. 11 also shows the ability of the user to change the width andheight of the spectra images 2006. FIG. 12 shows the output of thesoftware with the spectra grouped by similarity into twelve families.

The entire recrystallization process was repeated with a different glassplate in the crystallizer, this time dispensing the hot solutions forrecrystallization directly into the crystallizer subassembly (shown inFIG. 21, as described herein) in the same well format. The crystallizersubassembly was sealed and placed in a Torrey Pines incubator at 70° C.The temperature was decreased to 10° C. over an eight hour time period.After sitting at 10° C. overnight, aliquots of the mother liquor wereremoved and diluted with acetonitrile to determine the solubility ofphenylbutazone in the various solvents at 10° C. by LC analysis. Theremainders of the solvents were removed by pipette and any residualsolvent was removed by wicking with filter paper. Disassembly of thecrystallizer gave crystals in array format, with each crystal in aseparate region on a flat glass substrate. The glass substrate was thenmounted vertically on an X-ray diffraction machine, as described aboveand data was acquired on selected elements. Comparison of the XRD twotheta plots clearly shows the presence of three polymorphs, shown inFIG. 10.

Example 4 Cimetidine—Polymorph Study

A solution of cimetidine in dichloromethane (25 mg/mL) was dispensedinto wells of a microtiter plate having removable vials that were 750 μlin size such that each well contained 10 mg of phenylbutazone. Afterremoval of the solvent by evaporation, sixteen recrystallizationsolvents were dispensed as described above in Example 3 and in Table 6,above. The reactor was sealed and heated to 60° C. for four hours. Thecover was removed and aliquots (e.g., 200 μl) of individual samples wereremoved by pipette and dispensed to the crystallization assembly (shownin FIG. 21, as described herein) in the same well format. Anotheraliquot was removed and added to an array of vials and diluted withacetonitrile for further dilution to determine the solubility ofcimetidine in the various solvents at 70° C. by LC analysis. Thecrystallizer assembly was sealed and placed in a Torrey Pines incubatorat 70° C. The temperature was ramped over 8 hours to 10° C. Aftersitting at 10° C. overnight, aliquots of the mother liquor were removedand diluted with acetonitrile for LC analysis. The solvents were removedby pipette and residual solvent was removed by wicking with filter.Removing the cover and disassembly of the crystallizer gave crystals ona flat glass substrate.

The glass plate was then placed between cross polarizing filters andscanned to obtain dry birefringence and birefringence images of selectedwells were obtained under a microscope equipped with crossed polarizedfilter. Raman spectra were obtained on individual crystals in each ofthe wells as described in the previous example.

Example 5 Sample Code for Polymorph Characterization

Various programming languages can be implemented to build software thatcan perform categorization of crystalline structures in accordance withthe principles of the present invention. Listing 1, below, illustrates aportion of Java® code that can be used to implement the categorizationprocess described in conjunction with FIG. 8.

Listing 1 package com.symyx.webapp.projects.polymorphs; importcom.symyx.webapp.xydata.filereader.*; import com.symyx.webapp.xydata.*;import java.util.*; import java.io.*; public classSignalProcessorPolymorphs { private SignalProcessorResultPolymorphsresult_;    public SignalProcessorResultPolymorphs    result( ) { returnresult_; }    public void    result(SignaIProcessorResultPolymorphs   theResult) { result_=    theResult; }    privateSignaIProcessorParametersPolymorphs    parameters_;    publicSignaIProcessorParametersPolymorphs    parameters( ) { returnparameters_; }    public void   parameters(SignaIProcessorParametersPolymorphs    params) {   parameters_= params; } public Signal ProcessorPolymorphs( ) {   this. initialize( ); }    private void initialize( ) {     this.result(new      SignalProcessorResuItPolymorphs( ));     this.parameters(new      SignalProcessorParametersPolymorphs( ));   } public void sort(Vector items, Vector baskets) {    if (items ==null) {      return;    }    int itemCount = 0;    float cc;    int i=0;    int j = 0;    XyDataSet theSet = null;    XyDataSet[ ] xySets =new XyDataSet[2];    Vector stdltems = null;    Vector stdBaskets =null;    Vector ccScores = new Vector( );    Float score = null;   Float topScore = null;    int scorelndex = 0;    int topScorelndex =0;    Vector. theForm = null;    int formCount = 0;    float minCC =this.parameters( ).minGroupCC( );    boolean useFixedReference =   this.parameters( ).useFixedReference( );    // standardize items tobe sorted    itemCount = items.size( );    stditems =this.standardize(items);    //System.out.println(“made stditems”);    //standardize the known forms    if (baskets != null) { // input not empty   stdBaskets = this.standardize(baskets);    }    else {    // no knownforms yet,    // so create new vector and put the first    item in //there    stdBaskets = new Vector( );   stdBaskets.add(stditems.elementAt(0)); } //System.out.println(“madestdBaskets”); // make form baskets for (j=0; j<stdBaskets.size( ); j++){    theForm = this.result( ).addNewForm( );   theForm.add(stdBaskets.elementAt(j)); } for (i=0; i<itemCount; i++) {// clean out the old data ccScores.clear( ); // get the next item to beclassified theSet = (XyDataSet)stdltems.elementAt(i); xySets[0] =theSet; //System.out.printin(“sorting item: ” + i + “ ” + theSet);//System.out.println(“xySets[0]: ” + xySets[0]); // get the formsformCount = this.result( ).formCount( );//System.out.println(“formCount=” + formCount); for (j=0; j<formCount;j++) {    xySets[1] =    (XyDataSet)this.result( ).getFormData(j,   useFixedReference);    //System.out.println(“xySets[1]: ” +   xySets[1]);    // align the data sets    XyDataMath.alignX(xySets);   //System.out.println(“xySets[0]: ” +    xySets[0]);   //System.out.println(“xySets[1]: ” +    xySets[1]);    // runcorrelation    cc = XyDataMath.corrcoef(xySets[0],    xySets[1]);   xySets[1].fitness(cc);    //System.out.println(“cc=” + cc);    // addto the vector    ccScores.add(j, new Float(cc)); } // find out which isthe best topScorelndex = 0; topScore =(Float)ccScores.elementAt(topScorelndex); for (j=0; j<formCount; j++)   score = (Float)ccScores.elementAt(j);      if (topScore.floatValue( )<      score.floatValue ( )){         topScore = score;        topScorelndex = j;      }     //System.out.println(“topScorelndex=” +      topScorelndex); }//System. out. println(“top score: ” //+ topScore.floatValue ( )); //qualified for one of the classified forms if(Math.abs(topScore.floatValue( )−1.0f) < java.lang.Float.MIN VALUE) {continue; // the same data set } else if (topScore.floatValue( ) >=minCC) {    theForm =    this.result( ).getForm(topScorelndex);   theForm.add(theSet);    //System.out.println(“old Form”); } else { //found a new form    theForm = this.result( ).addNewForm( );   theForm.add(theSet);    //System.out.println(“newForm added”); } } }/**  * standardize the data sets by removing the slope from the    datasets,  * and perform normalization on them  */ private Vectorstandardize(Vector theSets) {    if (theSets == null) {    return null;   }    Vector stdSets = new Vector( );    XyDataSet theSet = null;   String fileName = null;    int count = theSets.size( );    for (inti=0; i<count; i++) {      //System.out.println(“i:” + i);      theSet =      (XyDataSet)theSets.elementAt(i);     //System.out.println(“theSet:” +      //theSet.header( ).getPath());      fileName =      theSet.header( ).getDataName( );      theSet =XyDataMath.removeSlope(theSet);      theSet =XyDataMath.normalize(theSet);      theSet.header().setDataName(fileName);      stdSets.add(theSet);    }      returnstdSets;    }    public Vector loadData( ) {      String baseDir =     “D:/raman_data/sorted_forms/”;      String listFileName =“Z_all_forms.txt”;      String fileName = null;      String filePath =null;      Vector fv = new Vector( );      TextFileReader dataReader =new      TextFileReaderPolymorphsRaman( );      filePath = baseDir +IistFileName;      FileReader fileReader = null;      LineNumberReaderreader = null;    try {      fileReader = new FileReader(filePath);     reader = new      LineNumberReader(fileReader);    }    catch(IOException e) {      e.printStackTrace( );    }    XyDataSet xyData =null;    int i=0;    try {    while ((fileName = reader.readLine( )) !=   null) {      try {         if (fileName.indexOf(“%”) >= 0) {          continue;      }         i++;         filePath = baseDir +fileName;         System.out.println(“loading... ”         +.filePath);        xyData = new XyDataSet( );         dataReader.readFile(filePath,     xyData);         fv.add(xyData);        //System.out.println(xyData.header(      ).getDa        //taName( ));    }catch (Exception e) {      e.printStackTrace();    }    } } catch (Exception e1) {    e1.printStackTrace( ); }   finally {      if (reader != null) {         try {        reader.close( );           }catch (IOException e) {         }     }    }    return fv;    }    public static void main(String[ ]args) {      SignaIProcessorPolymorphs spp =      newSignaIProcessorPolymorphs( );      if (args.length > 0) {        spp.parameters( ).minGroupCC(java.lang.Float        .parseFloat(args[0]));         spp.parameters().useFixedReference(true)         ;      }    Vector unknowns =spp.loadData( );    System.out.println(“..........”);   System.out.println(“sorting...”);    spp.sort(unknowns, null);   spp.result( ).printForms( );   } }

Example 6 Crystallization Workflow

An exemplary crystallization workflow is shown in FIG. 31. In theworkflow, the crystallization solvents 3101 and precipitation solvents3103 of interest are placed in solvent rack 3105 for use in theworkflow. Array 3110 comprising the drug candidate of interest (a saltor a neutral compound) from the reaction station is placed in reactorassembly 3120. The array can be an 8×12 array (e.g., a 96-well plate) orany other array known in the art (e.g., a 384-well plate).Crystallization solvents from the solvent rack are deposited into thewells of the array and the array is equilibrated to at least partiallydissolve the drug candidate in the solvent. In one embodiment, 800 μL ofthe crystallization solvents are added to each well, although any volumeof solvent may be used in accordance with this invention.

After the drug candidates have been equilibrated with thecrystallization solvents to form solution, the solutions are filtered infiltration assembly 3130. In one non-limiting embodiment, 650 μL istaken from each well of the reactor assembly and filtered in thefiltration assembly and a 600 μL aliquot of each sample is removed afterfiltration. The aliquots may then be daughtered for crystallizationanalysis and other analyses into one or more different apparatus. In oneembodiment, 50 μL of each sample is aliquoted into liquid chromatography(LC) vials 3140 for determination of solubility at a high temperature;250 μL from each sample is aliquoted into crystallizer assembly 3145 toinvestigate crystallization by cooling; 200 μL from each sample isaliquoted into evaporation plate 3150 to investigate crystallization byevaporation; and 100 μL of each sample is aliquoted into precipitationplate 3155 along with 400 μL of a precipitation solvent to investigatecrystallization by precipitation.

After crystallization, supernatants from the samples subjected tocrystallization in the crystallizer assembly and precipitation plate arecollected. The supernatant from the crystallization assembly may bealiquoted into LC vials 3165 for determination of the solubility of thedrug candidate at a cold temperature (e.g., room temperature or below).The supernatant may be collected at station 3160 for any purposedesired, such as recycling or discarding the drug candidate.

One having ordinary skill in the art following the teachings of thisinvention would recognize that one could use different combinations ofcrystallization apparatus and/or methods. For instance, one couldprovide a number of crystallizer assemblies that use differentcrystallization temperatures, or could use crystallizer assemblies toperform precipitation or evaporation crystallization. Further, onehaving ordinary skill in the art would recognize that one could performdifferent non-crystallization analyses using this workflow, such as logP analyses.

All publications and patent applications cited in this specification areherein incorporated by reference as if each individual publication orpatent application were specifically and individually indicated to beincorporated by reference. Although the foregoing invention has beendescribed in some detail by way of illustration and example for purposesof clarity of understanding, it will be readily apparent to those ofordinary skill in the art in light of the teachings of this inventionthat certain changes and modifications may be made thereto withoutdeparting from the spirit or scope of the appended claims.

1. A needle assembly for aspirating a liquid out of or dispensing liquidinto a sealed receptacle, the assembly comprising: a first body; atleast one guide member supported by the first body for sliding movementbetween an extended position and a retracted position; a second bodyattached to said at least one guide member, the second body beingrelatively farther from the first body when the guide member is in theextended position and relatively closer to the first body when the guidemember is in the retracted position; a biasing member biasing the guidemember to the extended position; and a liquid handling needle coupled tosaid first body so sliding movement of the guide member toward theretracted position retracts the second body relative to the liquidhandling needle, said second body being adapted to contact the sealedreceptacle during insertion of the liquid handling needle into thereceptacle and during removal of the liquid handling needle from thereceptacle, said biasing member urging said at least one guide membertoward said extended position during said insertion and during saidremoval whereby the second body remains in contact with the sealedreceptacle as the guide member moves from its extended position to itsretracted position during said insertion and from its retracted positionto its extended position during said removal.
 2. A needle assembly asset forth in claim 1 wherein the second body moves in conjunction withsaid at least one guide member.
 3. A needle assembly as set forth inclaim 1 wherein the biasing member comprises a spring.
 4. A needleassembly as set forth in claim 1 wherein the spring is retained betweenthe first and second bodies by the guide.
 5. A needle assembly as setforth in claim 1 wherein the liquid handling needle extends into thefirst body and is connected to a mechanism operable to transfer a liquidthrough the liquid handling needle.
 6. A needle assembly as set forth inclaim 1 wherein the at least one guide member comprises a first guidemember and a second guide member, the first and second guide members areeach supported by the first body for sliding in parallel relative to thefirst body, and the second body is attached to each of the first andsecond guide members so the second body moves with the first and secondguide members.
 7. A needle assembly as set forth in claim 1 wherein theliquid handling needle extends a fixed distance from the first body. 8.A needle assembly as set forth in claim 1 further comprising a ventingneedle supported by the second body.
 9. A needle assembly as set forthin claim 8 wherein the venting needle is held in a fixed positionrelative to the second body and the liquid handling needle is held infixed position relative to the first body.
 10. A needle assembly as setforth in claim 8 in combination with a robot having a moveable arm, theneedle assembly being mounted on the arm for movement of the needleassembly by the robot.
 11. A needle assembly as set forth in claim 1 incombination with a robot having a moveable arm, the needle assemblybeing mounted on the arm for movement of the needle assembly by therobot.
 12. A needle assembly as set forth in claim 1 wherein the secondbody comprises a guide block.
 13. A needle assembly as set forth inclaim 1 wherein the guide member comprises a rod.